Lecture 17 - Stem Cells and their use in therapy

Question 1

What is the teratoma assay?

Answer 1

way of determining pluripotency
Teratomas are benign cancers that have tissues within them from all 3 germ layers. Take a population of pluripotent stem cells, its defining criteria are that it can differentiate into cells from all 3 germ layers
Inject into immunocompromised mouse subcutaneously - if truly immunocompromised should give rise after a few weeks to a teratoma, that can be analysed histologically. Different structures can be identified that can be linked to different germ layers. Test of stem cell functionality is to cause a defect and see if stem cell can restore functionality

Question 2

Describe transcriptional control of ESCs

Answer 2

Pluripotent transcription factors:
Oct4, Nanog, Sox2.

Act collectively to regulated ESC pluripotency
These transcription factors expressed very early on in embryonic stem cells - activate pluripotency associated pathways and suppress pathways required to move cell on towards differentiation. As cells start to differentiate they lose expression of Oc4, Nanog and Sox2

ES cells have the ability to self-renew or differentiate into cells of all three germ layers (endoderm, ectoderm, and mesoderm). In ES cells, OCT4, SOX2, and NANOG form a core transcriptional network influencing the stem cell self-renewal machinery.

Question 3

Give examples of the therapeutic potential of ESCs

Answer 3

Generation of specific cell types and tissue structures to replace worn or diseased body parts, for example:
Neurodegenerative diseases
Diabetes
Corneal defects
Cardiovascular disease
Musculoskeletal disorders
Other Applications:
Toxicity testing - drug testing
Model systems - engineering tissues could replace animal testing

Question 4

Describe potential difficulties of ESC therapies

Answer 4

Important to get cells to differentiate fully to form a functional tissue. If you fail to do it fully and a pluripotent stem cell still present in therapeutic tissue this could give rise to teratoma causing a tumour

Question 5

Describe adult stem cells

Answer 5

Multipotent:

Stem cells that can differentiate into more than one cell type but have a more restricted differentiation potential than pluripotent stem cells.

More specifically, adult multipotent stem cells generally have the ability to differentiate into the cells of the tissues in which they reside.

Question 6

Describe the therapeutic potential of adult stem cells

Answer 6

Allows use of autologous cells
Fewer ethical concerns?
Fewer safety concerns? Doesn’t give rise to teratomas
Examples:
Haematopoietic stem cells
Mesenchymal stem cells
Epidermal stem cells
Neural stem cells
Limbal (corneal) stem cells

Question 7

What are induced pluripotent stem cells?

Answer 7

The introduction of genes associated with embryonic stem (ES) cell pluripotency into somatic cells, such as fibroblasts, can reprogramme somatic cells to pluripotent stem cells.
The introduction of just four (or fewer!) factors appears to be sufficient:
Oct-4, Sox2, Nanog and Lin28
Oct-4, Sox2, Klf4 and c-Myc

Question 8

What are the issues with iPS cells?

Answer 8

Reprogramming is inefficient (typically <1%)

• Depends on:
  a) The number of cells that take up the virus/vector carrying the reprogramming factors (viral transduction efficiency) and
  b) The number of virally transduced cells that actually undergo reprogramming.

Use of viral delivery systems
Use of oncogenes (cMyc) cancer risk
Retain epigenetic memory of parent cells (unlike SCNT)
Teratoma risk