Cloning and Stem Cells

A wood panel painting (~1500A.D.) depicting the limb transplantation miracle by Saints Cosmos and Damian.  According to legend Cosmos and Damian were physicians who successfully transplanted the limb of a dead Moor onto a patient whose leg required amputation.

• Red Deer replace their antlers every year. It takes ~3 months.• We replace our blood cells. ~100 million /hour• If part of the human liver is removed, it will re-grow• Salamanders and Newts can re-grow their limbs

•Video – newt limb regeneration

Can we regenerate parts of the body?

In general:

regeneration in vertebrates doesn’t work very well

Limbs and most organs aren’t replaced

Can cells be reprogrammed?

Experimental Example: Amphibian tadpole:Preprogrammed Muscle cells were transplanted into gut region of cells in the embryo but they stayed as muscle cells.

Cells are programmed and “know’ what they will become if the cells stay intact. How?

Nucleus expresses genes→ makes RNA → RNA makes proteins. Some of that protein goes back into the nucleus and programs the cells to keep making more of their kind.

What is a Stem cell?

• Can reproduce itselfOr

• Can enter a differentiation pathway whereby one of its daughters is differentiated and the other stays an undifferentiated stem cell (totipotent)

Where do we get stem cells?

A reliable stem cell reference: http://www.isscr.org/science/faq.htm

Stem cells can be found naturally in 1) A Fertilized Egg (embryonic stem cells (ES)):

ES are Also found in small numbers in various tissues in the fetal and adult bodyExamples:

• blood stem cells are found in bone marrow → give rise to all specialized blood cell types. • newborn baby: umbilical cord, possibly baby teeth and amniotic fluid

Video: (human emb. development)

150 cells called (blastocyst) contains two types of cells: I) trophoblast II) inner cell mass

Embryonic stem cells are obtained from the inner cell mass.

2. Cancers can form a variety of cell types(connective tissue, cartilage, blood cells) can be transplanted into host animals and in some cases turn into non-cancerous normal cells

3. (Since 1981) Start w/ an embryo.Blastocyst cells can be put in culture and

grow indefinitely. When put into a host they will differentiate

into a variety of cells.

Other ways to get stem cells:

Ovarian cancer stem cells

Stem cells tend to make a heterogeneous collection of differentiated cells

Video: creating lines

We haven’t figured out how to make stems cells differentiate into every single type of tissue. …..Yet.

Advantages of Embryonic Stem cells

• Capacity for indefinite proliferation in culture

• Can be differentiated into a variety of cells including germ cells

• Can be maintained as frozen stocks

•Problems:–Undergo spontaneous genetic changes–Hard to make into certain cell types (e.g. pancreas) (nerve tissue is fairly easy to make, however)–Immuno-rejection unless patient is immuno-suppressed. (not ideal).

Nuclear reprogramming and Therapeutic cloning

• Can we make rejuvenated embryonic cells from adult differentiated cells (of the same individual)?

• This would avoid the problem of immuno-rejection

Somatic Cell Nuclear transfer

1. Start with egg (frog, mouse, human)2. Use UV to kill pronucleus (egg chromosome)-left

w/ cytoplasm3. Pick up another cell in a way such that the cell

wall is broken but intact nucleus (using a special micropipette)

4. Inject the intact nucleus into the cytoplasm-only egg cell. The cell thinks It’s been fertilized. It starts to grow into an embryo and then an adult.

http://www.dnalc.org/cloning.html

•This cloning is inappropriate in humans (called reproductive cloning) video: SCNT

Problems w/ SCNT (somatic cell nuclear transfer)

• Some transplanted nuclei show a long-term memory of their original gene state so they won’t allow reprogramming.

• only about 1/3 of nuclear transplants work

However, we’re making progress!

Legal Issues

In the United States, as of April, 2009:

• it is illegal to use federal funding for research on embryos derived from SCNT.

• It is legal to use federal funding for research on embryos derived from IVF, if those embryos are not going to be used for anything else (like making a baby).

Long-term GoalReprogram cells without having them go

through a nuclear transfer to an egg

Sept. 2007 “Researchers Isolate Adult Stem Cells for First Time in Tendon tissue” (adult fibroblasts)

Maybe a cure for Alzheimer's disease, Parkinson's disease, and spinal cord injury,

In the news:

Lots of stories out there….

Induced Plueripotent Stem cells (iPS) also called Adult Stem cells

iPS cells from adult human cellsDeveloped by two independent research teams:

James Thomson and colleagues at University of Wisconsin-Madison

Shinya Yamanaka and colleagues at Kyoto University, Japan

(1)Isolate and culture donor cells.

(2)Transfect stem cell-associated genes into the cells by viral vectors. Red cells are successes!

(3)Harvest and culture cells according to ES cell culture, using mitotically inactivated feeder cells (lightgray)

(4)A small subset of the transfected cells become iPS cells and generate ES-like colonies.

5 Common Ethical Objections• Slippery slope – where do you draw the line

between frogs & mice, and humans• Cloning is unnatural. (Antibiotics are unnatural

also)

• Killing of a potential life. “An embryo = a human being” Actually, an embryo would never

progress until it is put into a womb.• God intends humans to suffer – we need to

have diseases.• It’s good for us to see how others tolerate suffering.