Lecture 25: Stem cells, Regenerative medicine

Question 1

What are stem cells

Answer 1

Unspecialised cells (primitive) that are capable of self renewal by cell division with potential to develop into many different cell types in the body

Question 2

Why are stem cells important

Answer 2

• Can repair and replace lost tissues

- Can be induced to become tissue specific ‘mature’ cells with special functions

Question 3

What is the difference in potency of adult stem cells vs embryonic stem cells

Answer 3

Adult: Multipotent or unipotent - can differentiate but to only one (skin stem cell) or few lineages (haemopoietic stem cell)
Embryonic stem cells: pluripotent- give rise to any cell in the human body from the 3 germ layers

Question 4

What is totipotent cell

Answer 4

cells that can give rise to embryo + extra embryonic tissue ie placenta

Question 5

What is a mesenchymal stem cell - multipotent

Answer 5

• Traditionally found in bone marrow but can be isolated for other tissues
• Can turn into fat, cartilage, bone, muscle, tendons and skin.
• Releases anti inflammatory and immunomodulatory factors

Question 6

What are all the organs that have adult stem cells

Answer 6

blood, gut, skin, brain, eye, muscle, mesenchymal stem cell- so not all organs have them

Question 7

What are some adult stem cell therapies

Answer 7

Stem cells in transplants:

bone marrow transplant, skin grafts and corneal transplant

Question 8

What are the two types of pluripotent stem cells (not found in vivo but share characteristics with cells in the early embryo

Answer 8

• Embryonic stem cells (ES) derived from the inner cell mass of the embryo
• Induce pluripotent stem cells (iPSC) from normal cells reprogrammed

Question 9

How is the pluripotency of Inner cell mass cells tested

Answer 9

Take ICM cells from one organism into another’s blastocyst to see the contribution of the stem cells to the individual.

• Tetraploid complementation : This is making a tetraploid blastocyst host and injecting the ES into that, so if you get a whole mouse then its from the ES
• ES cells are injected under the kidney capsule of a mouse to check for a teratoma - tumour with 3 germ lines.
• Embryoid body assay: inducing differentiation of hES into embryoid bodies using growth factors then into a further tissue which can be tested for the different germ layers
• immunostaining for stem cell markers or transcription factors

Question 10

How are Embryonic stem cells derived

Answer 10

The inner cell mass is isolated from the cultured blastocyst. This is cultured in vitro from which grows colonies of stem cells. They can be maintained indefinitely in cell culture because they keep dividing.

Question 11

Human stem cells … be injected into mouse blastocyst

Answer 11

no -ethics

Question 12

What test is used to check for the pluripotency of human cells

Answer 12

Teratoma assay (kidney mouse tumour), immune staining of stem cells, embryoid body assay

Question 13

What are the possible uses for human ES cells

Answer 13

• Tissue engineering
• Cell based therapies- maturing, purifying and transplanting stem cells
• Models for long term diseases

Question 14

What are the dangers of ES being used in therapies

Answer 14

• they can develop into teratomas if there are stray undifferentiated ES left
• Ethical considerations for destruction of the embryo
• Have to avoid rejection so choose immunological privilegd.

Question 15

What are the differences between normal tissue and ES cell derived tissues

Answer 15

There are some minor epigenetic differences so differences in amount of expression of the gene but it will be in the right place and be on

Question 16

What is reprogramming done by somatic cell nuclear transfer

Answer 16

This is replacing the nucleus of oocyte with the nucleus of an already differentiated cell. This works because there are factors oct4, sox2, klf4 and cMyc that reprogram the somatic nucleus back to the embryonic state

Question 17

What is the difference between reproductive cloning and therapeutic cloning

Answer 17

From somatic cell nuclear transfer the is formed an embryo: taken to term is reproductive cloning but therapeutic cloning is harvesting the ES cells for therapy

Question 18

What is the difference between induced pluripotent stem cells

Answer 18

ES line is gold standard, with differences in methylation and expression levels different but IPs more moral and no immune rejection but still risk of teratoma

Question 19

How are iPS cells used for gene editing

Answer 19

The IPs cell line is taken from a patient with genetic defect, and these cells can be edited using crispr and then differentiated into the right cell and reintroduced to the patient.
- animal’s gene for creating an organ is knocked out in embyro and then the hiPS cells injected into the blastocyst to grow the human organ in the animal

Question 20

What is an organoid- used to bridge 2D cell culture and in vivo study

Answer 20

A mini 3D structure 200-500 cells derived from either pluripotent stem cells or adult stem cells or progenitors, in which cells spontaneously self- organise into functional cell types and recapitulate some function of the organ.
- Can be used to study organ function from genetic disease