lecture 9 - stem cells Flashcards
What is a stem cell?
Two defining criteria:
Stem cells have the ability to self-renew
…by asymmetric cell division (gives rise to one stem cell and one daughter cell destined for differentiation.
Stem cells give rise (differentiate) to different specialised cells types (blood cells, skin cells, bone cells)
Describe the changes in potency of stem cells
day 0 - fertilisation
day 3-4 - totipotent cells - brief stage before blastocyst formed
day 5-8 - blastocyst - inner cell mass pluripotent cells which have ability to go on and give rise to all cells, go on to form embryo
What are totipotent stem cells?
Total Potential
Have the ability to develop into an entire organism
The fertilised egg
Daughter cells up to around Day 4 following fertilisation and before blastocyst formation
What are pluripotent stem cells?
Several/Many Potentials
Have the ability to develop into virtually every cell type
Do not form placenta and supporting tissues needed for foetal development, unable to generate a new organism on their own, therefore not “totipotent”
ES cells of the inner cell mass (ICM) in the blastocyst are pluripotent
How do you grow embryonic stem cells in vitro?
Pluripotent cells from the ICM of the blastocyst separated from the surrounding trophectoderm.
Inner cell masses plated into culture dishes and grown in nutrient medium supplemented with serum, supported by irradiated fibroblast feeder layers.
Cells begin to divide over a period of approximately two weeks to form colonies
Colonies removed, dissociated and replated to allow further expansion
Cells continue to divide for several months without differentiating – allows expansion of pluripotent stem cell pool
How do you determine the pluripotency of ES cells?
sub-cutaneous injections of human embryonic stem cells into immunocompromised mice form teratomas
What are teratomas?
Teratomas contain differentiated cell types derived from all three germ layers:
Ectoderm
Mesoderm
Endoderm
In vivo method of determining pluripotency
Describe the transcriptional control of ES cells
Oct-4, Sox2 and Nanog = “pluripotency transcription factors”
Oct-4
Transcription factor expressed by embryonic stem cells
At the blastocyst stage, Oct-4 is only expressed by ES cells in the inner cell mass
Sox2
Transcription factor that forms a complex with Oct-4.
Expression pattern similar to Oct-4.
Nanog
Named after “Tir nan Og”, the Celtic land of the ever young
Transcription factor, expressed specifically by ES cells (ICM), slightly later than Oct-4
Oct-4, Sox2 and Nanog appear to function co-operatively in the maintenance of pluripotency and self-renewal of ES cells
Describe how ES cells use telomerase
Embryonic stem cells express high levels of telomerase, an enzyme that helps maintain the protective function of telomeres at the end of chromosomes
Little or no telomerase activity in mature differentiated cells
What are Induced pluripotent stem cells (iPSCs)?
have properties similar to embryonic stem cells
Oct4 and Sox2 artificially overexpressed to induce pluripotency, reverse development
What are adult stem cells?
rare.
Multipotent: Give rise to different cell types of the tissue in which they reside.
Usually generate an intermediate cell type before they fully differentiate, often termed a “precursor” or “progenitor” or “transit amplifying” cell.
Progenitor cells are “committed” to a specific differentiation pathway.
Properties:
Generally described as multipotent
Can self-renew for long periods of time
But appear to have less self-renewal ability than ES cells
Due, in part, to reduced telomerase levels
Differentiate into different cell types with specialised functions
Primarily function to maintain the steady-state activity of a cell and its resident tissue
May help replace cells that are lost through injury and/or disease
What are the self-preservation mechanisms of adult stem cells?
Long-lived, but slowly cycling
High ABC transporter expression (discards potentially harmful toxins)
High aldehyde dehydrogenase (ALDH) expression (converts acetaldehyde to acetic acid)
Give examples of adult stem cells
Haematopoietic stem cells
Blood: Neutrophils, erythrocytes, megakaryocytes
Epidermal stem cells
Skin: Keratinocytes, hair, gland cells
Mesenchymal stem cells
Bone: osteoblasts
Cartilage: chondrocytes
Adipose tissue: adipocytes
How are cell surface markers expressed?
Cell surface markers expressed by human haematopoietic stem cells (HSCs)
Cells express specific proteins that can act as a “fingerprint” guide to their identity
CD34+
CD133+
Thy1+
CD38-
Lin-
Antibodies targeted to one or more of these cell surface proteins can be used to identify and isolate HSCs
Where are HSCs located?
HSCs are located in specialised microenvironments in bone marrow:
The HSC Niche
Contact dependent or juxtacrine signalling
when it divides a transmit amplifying cell also produced
