Stem cells

Question 1

What is a stem cell?

Answer 1

• cells able to divide to give rise to new cells required to make and maintain the cells, tissues and organs of multicellular organisms.
• undifferentiated, not engaged in a specific physiological function
• have “potential”
• “self-renewal”

Question 2

Describe “potential”

Answer 2

• aka potency
• a population of stem cells can be the source of some, many or even all cell types in an organism

Question 3

Describe “self-renewal”

Answer 3

maintained through rounds of cell division

Question 4

Describe asymmetric cell division

Answer 4

gives rise to one “stem cell” and one “committed cell”

Question 5

Describe symmetric division with asymmetric population behaviour

Answer 5

gives rise either to two “stem cells”, OR two “committed cells”

Question 6

Cells acquire … during development

Answer 6

more restricted fates

Question 7

Describe development from the zygote

Answer 7

• blastula
• gastrula
• germ cells (male, female) and derm layers (endo, meso, ecto)

Question 8

Describe the endoderm

Answer 8

• stomach cells make digestive tube
• thyroid cells in pharynx
• alveolar cells in respiratory tubes
• intestinal epithelium cells

Question 9

Describe the mesoderm

Answer 9

• dorsal notochord
• paraxial bone tissue
• intermediate kidney tubule cells
• lateral erythrocytes, lymphocytes and monocytes
• facial muscles in the head

Question 10

Describe the ectoderm

Answer 10

• epidermal fibroblast on outer surface
• neurones of CNS
• melanocytes of neural crest

Question 11

Describe fate-mapping

Answer 11

identifies early cells

Question 12

Describe totipotency

Answer 12

• makes everything, including extraembryonic tissue (placenta)
• zygote and morula

Question 13

Describe pluripotency

Answer 13

• can make all cells in the embryo and adult
• germ layer specified
• blastocyst

Question 14

Describe multipotency

Answer 14

• all cell types of a tissue or organ
• germ layer specified
• blastocoel and grastula

Question 15

Describe oligopotency

Answer 15

can make several, related cell types

Question 16

Describe unipotentcy

Answer 16

can make only one cell type

Question 17

What happens at the grastula boundary?

Answer 17

• differentiation
• proliferation
• migration
• signalling
• morphogenesis
• organogenesis
• growth
• homeostasis
• regeneration

Question 18

Describe the lineage process

Answer 18

• stem cells become progressively more ‘determined’ during development
• potency deceases as lineage progresses

Question 19

Describe adult stem cells

Answer 19

continue to maintain our tissues and organs in response to ”wear and tear”, ”infection/disease” and
“other environmental factors/toxins”

Question 20

Describe developmental stem cells

Answer 20

maintained as adult stem cells

Question 21

Which stem cells makes intestinal epithelial cells?

Answer 21

intestinal stem cells

Question 22

Which stem cell makes lymphocytes, monocytes and erythrocytes?

Answer 22

haemtopoetic stem cells

Question 23

Which stem cells makes skin fibroblast?

Answer 23

epidermal stem cell

Question 24

Describe transit amplifying cells

Answer 24

• derived from stem cells later in development and adult stem cells
• lineage committed
• neither differentiated nor “self- renewing”
• divide several times to expand to a population of cells that eventually differentiates
• reduces the number of divisions a stem cell makes

Question 25

Describe gene regulation inside stem cells

Answer 25

- TFs - epigenetic/chromatin modifications - microRNAs - external signals can drive differentiation to the required fates - for control and manipulation of stem cells

Question 26

Describe TFs in stem cells

Answer 26

- maintain a stable undifferentiated state - supress genes that drive differentiation

Question 27

Describe epigenetic/chromatin modification in stem cells

Answer 27

- histone modifications - DNA methylation - transcriptionally silence regions that would lead to differentiation - allow expression of genes for the present state

Question 28

Describe microRNAs in stem cells

Answer 28

alternate splicing of transcripts

Question 29

ICM

Answer 29

inner cell mass

Question 30

Describe the production of mammalian pluripotent embryonic stem cells

Answer 30

- taken from the ICM of embryos - highly defined mix of media and growth/signalling factors - maintained in an undifferentiated state - different levels of readiness to differentiate

Question 31

Describe the different levels of readiness to differentiate

Answer 31

naive > formative > primed

Question 32

Describe mammalian pluripotent ESCs

Answer 32

- pluripotent - able to form teratomas - distinct gene expression profile, associated with animal germlines

Question 33

Describe an embryo

Answer 33

- trophectoderm - inner cell mass - primitive endoderm

Question 34

Human embryonic stem cells can

Answer 34

self-organise and pattern in culture

Question 35

Describe the in vivo evidence of ESC pluripotency

Answer 35

- take blastocyst from mouse with white fur - inject ESC from mouse with black fur to create a chimera - second generation you can get a mouse with black fur

Question 36

Summarise the in vivo evidence of ESC pluripotency

Answer 36

generation of donor genotype in the F2 confirms contribution to the chimeric germline

Question 37

Describe gene targeting for transgenic mice

Answer 37

target gene undergoes homologous recombination

Question 38

Describe random integration for transgenic mice

Answer 38

integration and insertion within a random gene

Question 39

Describe assaying pluripotency

Answer 39

- in vitro experiments study cell competence and regulation over time - in vivo reimplantation of ESCs back into embryos allows observation of their contribution - genetic manipulation of cells for transgenic mice

Question 40

Describe iPSCs

Answer 40

- 4 TFs responsible for reprogramming activity - form teratomas - can contribute to all 3 germ layers, and the germline

Question 41

Describe Gurdon's expts

Answer 41

- nuclear transfer in frogs - prove the genome is not diminished as a method of gene regulation during differentiation - challenged prevailing theory that cells fates were stable - frog egg cytoplasm could reprogram the transplanted nucleus of a differentiated cell

Question 42

Which are the 4 TFs necessary for reprogramming?

Answer 42

- Oct3/4 - Sox2 - Klf4 - c-Myc

Question 43

c-Myc

Answer 43

oncogene

Question 44

Describe transcriptional control in “primed” pluripotent cells

Answer 44

- levels of TFs vary stochastically - addition of BMP4 pushes cells with low Oct4 to extraembryonic fates

Question 45

Describe epigenetic/chromatin based regulation of primed human pluripotent stem cells

Answer 45

- genes required for pluripotency are ON - genes required for later differentiation are OFF - genes that might need to switch on soon are in a BIVALENT state

Question 46

Describe gene bivalence

Answer 46

ready to switch on or to be silenced depending on the initial differentiation decision made

Question 47

Describe haematopoietic stem cells (HSCs)

Answer 47

- relatively rare (40k-200K) - produce >200 billion RBCs alone - >2.3 million cells produced per second - different progenitors in the lineage with different potencies

Question 48

Where did the stem cell concept arise?

Answer 48

study of the bone marrow transplants into irradiated mice

Question 49

How are different HSCs identified

Answer 49

- creating antibodies to distinct combinations of cell surface markers - FACS allows them to be isolated from the rest of the population

Question 50

FACS

Answer 50

Fluorescence Activated Cells Sorting

Question 51

Which diseases and conditions are treated with HSCs?

Answer 51

- sickle cell anaemia - autoimmune diseases

Question 52

Describe adult intestinal stem cells

Answer 52

- Paneth cells at the base of the intestinal crypts

Question 53

What is one underlying cause of ageing?

Answer 53

the capacity of stem cells to replicate and maintain tissues is limited