Anita Hall - Stem cells

Question 1

Link between cell signalling and stem cells?

Answer 1

Question 2

What are the different stages in a cell’s life cycle?

Answer 2

Question 3

What are the defining features of a stem cell?

Answer 3

Question 4

What are the different types of stem cells?

Answer 4

Question 5

Where are embryonic stem cells found? What potency do they have?

Answer 5

Question 6

What are the three germ layers that a ES cell can differentiate into?

Answer 6

Question 7

How can ES cells be stably maintained and expanded in vitro?

Answer 7

Question 8

Ethical issues associated with working on ES cells?

Answer 8

Question 9

What did John Gurdon discover about stem cells in (1962)?

Answer 9

Question 10

What did Takahashi & Yamanaka (2006) discover?

Answer 10

Question 11

When creating IPSCs, how long does reprogramming take and do different combinations of TFs introduced influence the stem cells produced?

Answer 11

Question 12

Before you can use IPSC in academic research, what do you have to do?

Answer 12

Question 13

IPSC test - Outline what the teratoma test is?

Answer 13

Question 14

IPSC test - What is the embryoid body test?

Answer 14

Question 15

IPSC test - On a genomic level how can we check whether our IPSCs resemble ES cells?

Answer 15

Correct genes are active - Promoters and histones of pluripotency genes expressed (demethylated -active) and pro-differentiation genes are repressed (e.g. methylated) – epigenomic modifications

Question 16

IPSC test - Growth of an entire organism

Answer 16

Question 17

What is the main safety concern when working with IPSCs?

Answer 17

Question 18

What did Hou P. et al show with regards to pluripotent stem cell induction?

Answer 18

Show that pluripotent stem cells can be generated from mouse somatic cells at a frequency up to 0.2% (low) using a combination of seven small-molecule compounds that don’t integrate into the genome – one of these molecule acts via Wnt signalling

The chemically induced pluripotent stem cells resemble embryonic stem cells in terms of their…

a) Gene expression profiles
b) Epigenetic status
c) Potential for differentiation
d) Germline transmission

By using small molecules, exogenous “master genes” are dispensable for cell fate reprogramming (no need to introduce genes) –> This chemical reprogramming strategy has potential use in generating functional desirable cell types for clinical applications.

Question 19

What are some examples of in-vitro assays performed on stem cells?

Answer 19

Question 20

What are some examples of in-vivo assays performed on stem cells?

Answer 20

Question 21

What two techniques are used for stem cell harvesting?

Answer 21

Stem cells harvesting

Selective adhesion - some stem cells have different adhesion molecule which can be taken advantage of

FACS sorting - method for sorting a heterogeneous mixture of biological cells into two or more containers, one cell at a time, based upon the specific light scattering and fluorescent characteristics of each cell.

Question 22

What is Lineage tracing?

Answer 22

Lineage tracing: a powerful technique and useful to understand

Lineage tracing experiments aim to highlight the full progeny of a given cell/cell population through genetic tagging - tagging stem cells/daughter cell/trans-amplifying cell and its progeny in order to trace them

Question 23

How is Lineage tracing performed using Cre-Lox as an example

Answer 23

Question 24

Using the example 82 yr. old ALS patient - explain how Skin cells were converted to iPS cells? What did they check for - think about the intentions?

Answer 24

Question 25

82 yr. old ALS patient - What did they do to their IPSCs in order to drive differentiation into Motor nuerons?

Answer 25

Question 26

How did Yang et al. 2013 go about investigating motor nuerons from ALS patients?

Answer 26

Question 27

What did Marchetto et al., 2010 find out about Rett syndrome using patient derived IPSCs?

Answer 27

Question 28

In another more recent Rett iPSC study, what did Chin EWM et al., 2016 show?

Answer 28

Question 29

What did Williams Syndrome patient derived IPSCs show in-vitro?

Answer 29

Question 30

Summary - Outline how patient derived IPSCs are generated and used in modelling disease phenotypes?

Answer 30

Question 31

In order to create a desired differentiated cell, do we always have to go back to pluripotency?

Answer 31

Question 32

What does this figure show?

Answer 32

Question 33

What are Tissue-specific stem cells?

Answer 33

Question 34

How to find/identify tissue stem cells?

Answer 34

Question 35

What are the different 'controllable parameters' (things that can change depending on the circumstance) from a stem cell to the fully differentiated cell?

Answer 35

Question 36

What is the stem cell niche?

Answer 36

Question 37

Common features of a stem cell niche?

Answer 37

Question 38

What are transit amplifying (TA) cells?

Answer 38

Question 39

Outline the crypt structure in the small intestine

Answer 39

Question 40

Outline the role of Wnt and Shh signals in BMP4 (Bone morphogenetic protein 4) expression in the small intestine crypt

Answer 40

Question 41

Small intestine - outline the role of EphB in maintaining the organisation of cells in the small intestine

Answer 41

Question 42

What are the two types of Stem cells found in the small intestine crypt?

Answer 42

Question 43

What did Luo and Puigserver, Nature 2016 reveal about the role of high fat diet on intestine stem cells?

Answer 43

Question 44

What are organoids and how are they used in research?

Answer 44

Question 45

What are skeletal Satellite Cells?

Answer 45

Question 46

What does the attached image reveal about the Satellite stem cell niche?

Answer 46

Question 47

How are Satellite cells activated?

Answer 47

Question 48

How do satellite skeletal muscle stem cells remain quiescent until activated?

Answer 48

Question 49

Outline how p38 MAPK helps to regulate satellite cell activity?

Answer 49

Question 50

What observation was made by Cheung, TH et al with regards to miRNA-489 and the cell cycle?

Answer 50

Question 51

What experimental setup was used to demonstrate the role of miRNA in quiescent adult stem cells? What were the results?

Answer 51

Question 52

What experiment Rodgers JT at al., Nature, 2014 perform to show the existance of a new G0 state?

Answer 52

Question 53

Once satellite stem cells are activated, can they divide asymmetrically or symmetrically?

Answer 53

Question 54

What is Duchenne muscular dystrophy? What happens to satellite cells?

Answer 54

Question 55

What is a potential future therapy for tackling Duchenne muscular dystrophy?

Answer 55

Question 56

In what way do old stem cell niches drive disrupt muscle stem cells?

Answer 56

Question 57

What happens when geriatric muscle stem cells switch from quiescence into senescence?

Answer 57

Question 58

What is one way that was suggested by Brack et al., Science, 2007 to slow down the rate of satellite cell aging?

Answer 58

Question 59

What was one signalling molecule that was indentified as having an effect on the presence of non-myogenic cells (non-muslce)?

Answer 59

Question 60

What effect did 'young blood' have on the neural stem cell niche?

Answer 60

Question 61

Definition of stem cell?

Answer 61

**Stem cell** - Stem cells are cells that have the ability to differentiate into specific cell types. The two defining characteristics of a stem cell are **perpetual self-renewal** and the ability to differentiate into a specialized adult cell type, the range of which is determined by the cell’s **potency**.

Question 62

Definition of self-renewal & Potency?

Answer 62

**Self-renewal** - The cell that has the ability to divide and produce progeny that is exactly the same as the originating/parental cell – non-differentiated. This bears a resemblance to cancer cells but stem cell division is highly regulated in contrast to the uncontrolled proliferation in cancer cells. **Potency** - The number of cell types that a stem cell can differentiate into Note - When approaching defining key words try to include… Definition Some molecular biology to help explain Examples

Question 63

What are two main advantages of using IPSCs over ES cells?

Answer 63

a) No need for embryo (ethical) b) Contain genetic information of the patient - useful for studying disease + reduced risk of rejection upon re-introduction

Question 64

Difference between neuronal and neural stem cells?

Answer 64

Neuronal - only produce neurons Neural - produce neurons + other CNS cells

Question 65

Outline the general steps in CNS development?

Answer 65

Question 66

Outline the process by which the neural tube is formed?

Answer 66

Question 67

Definition of Neural Stem cells?

Answer 67

**Neural stem cells** = stem cells of the nervous system that generate neurons and oligodendrocytes and astrocytes in the embryo and the adult CNS They are a type of ‘tissue-specific’ stem cells (called adult stem cells when not in the embryo)

Question 68

On a cellular level, explain what changes are occuring to the neural stem cells during development

Answer 68

Question 69

On a genomic level, outline what changes occur when a neuronal cell gives rise to glial cells?

Answer 69

Question 70

Are we able to make cerebral organoids?

Answer 70

YEAHHHH BUDDYYY

Question 71

Can postnatal and adult mammalian brains make new neurons?

Answer 71

Question 72

What is the brain's ventricle system?

Answer 72

The ventricular system is a **set of four interconnected cavities known as ventricles** in the brain. Within each ventricle is a region of choroid plexus which produces the circulating cerebrospinal fluid (CSF). Ventricular system and the central canal of the spinal cord are **lined with ependyma**, **a specialised form of epithelium** connected by tight junctions that make up the blood–cerebrospinal fluid barrier

Question 73

What are the neural stem cells of the subventricular zone?

Answer 73

Question 74

In rodents what are the progeny of the neural stem cells of the SVZ?

Answer 74

Question 75

How does the SVZ stem cell fate differ between humans and rodents?

Answer 75

Question 76

Is there a transcription factor programme that drives SVZ neural stem cell differentiation?

Answer 76

Question 77

Why do rodents have the NSCs in the SVZ?

Answer 77

Question 78

In rodents, what were the neural stem cells in subgranular zone (SGZ) responsible for?

Answer 78

Question 79

What are the characterisitics of Neural stem cells of the subgranular zone (SGZ) and what type of cells do they produce?

Answer 79

Neural stem cells of the subgranular zone (SGZ) of the dentate gyrus (hc) **Neural stem cells in SGZ** = a type of astrocyte/radial glia-like cell (**GFAP+, nestin+** (found on immature neural cells) and **Sox2+** (associated with neural/fated to be neural)) Quiescent, can self-renew & generate radial-glia like cells, neurons and astrocytes (some say subtypes have varying potency) The neurons formed are glutamatergic dentate granule cells

Question 80

What factors contribute towards the final cell fate of SGZ neural stem cells?

Answer 80

Question 81

Is there any evidence for neurogenesis in the human adult hippocampus SGZ?

Answer 81

Question 82

What functions do adult neurons generated from SGZ have?

Answer 82

Question 83

What regulates Adult NSC proliferation?

Answer 83

Neuronal activity nearby can stop SGZ NSC proliferation

Question 84

Does the vascular system help maintain the neural stem cell niche?

Answer 84

Question 85

What are some characterisitcs of aging (and/or diseased?) brains when it comes to NSC & neurogenesis?

Answer 85

Question 86

What is the cancer stem cell hypothesis?

Answer 86

Question 87

What is Glioblastoma Multiforme (GBM)?

Answer 87

Question 88

How could we use mesenchymal stem cells to target tumours?

Answer 88

A glioma is a type of tumor that starts in the glial cells of the brain or the spine

Question 89

Has using neural stem cells (iPSC) to treat dopaminergic neuron loss in Parkinsons disease been succesful?

Answer 89

Question 90

Where does Dopaminergic neuron development take place?

Answer 90

Question 91

What is the role of the Isthmus in dopaminergic neuron production?

Answer 91

Question 92

What signals do DA neurons in the Tegmentum receive during development?

Answer 92

Question 93

What are the two major Dopaminergic neuron projections in the brain?

Answer 93

Question 94

What happens during early DA development in the neural tube?

Answer 94

Question 95

What key TFs are associated with driving progenitor cells to form DA neurons?

Answer 95

1. Lmx1a 2. Msx1

Question 96

What are some NSC strategies to help treat and understand Parkinsons diseases? What are some key questions to take into consideration when using neural stem cells to treat Parkinson’s disease?

Answer 96

**One strategy: Replace diseased dopaminergic neurons** 1. How to prevent tumours from formed after transplantation? 2. What specific cell do we transplant – neuroblasts (more resilient – trouble differentiating) or DA neurons (less resilient/more fragile – already specialized) **Another NSC based strategy: neuroprotection (transplant supporting cells)** Neuroprotection - provide cell survival factors to allow them to survive for longer **Another use of NSCs in PD research** Modelling the disease in order to understand why dopaminergic neurons die in PD