Module 4 - Stem cells Flashcards
Endogenous stem cells
maintain human body
What is a stem cell?
A cell that is able to self renew and differentiate into mature functional cell types
Stem cell > Progenitor > Mature cell
Name 3 ways of differentiation
Asymmetric cell division
Symmetric cell division
Mixed model
Explain the asymmetric stem cell division models
A) Asymmetric localisation of cell polarity regulators initiates the asymmetric division
B) Segregation of cell fate determinants ot the cytoplasm of one daughter cell
C) Exit of the daughter cell from a stem cell niche triggers differentiation
Different functional outcomes of stem ells
Symmetric self-renewal: proliferation and maintenance of developmental potential = increase in stem cell pool, no generation of differentiated progeny
Asymmetric self-renewal: proliferation and maintenance of developmental potential = maintenance of stem cell pool, generation of differentiated progeny
Lack of self-renewal: proliferation only = depletion of stem cell pool, generation of differentiated progeny
Lack of self-renewal: No proliferation, maintenance only = maintenance of stem cell pool, no generation of differentiated progeny
Definitions of stem cell potency - Totipotent
can produce an entire organism i.e. zygote (egg) and extra-embrynonic (placenta)
Pluripotent
gives rise to cell derivatives of all 3 germ layers i.e embryonic stem (ES) cells
Multipotent
gives rise to multiple cell types e.g. haematopoietic SCs, Mesenchymal SCs
Unipotent
Gives rise to only one cell type (precursor) e.g. spermatogonial stem cells
Adult stem cells
Organ or tissue specific
- Asymmetric or mixed cell division
- multipotential and limited in vitro expression
- e.g. HSC in bone marrow, epidermal SCs, MSCs
Embryonic stem cells
- Symmetrical cell division
- pluripotent, can be cultured indefinitely
- ES derived from inner cell mass of blastocyst
- embryonic germ (EG) cells derived from primordial germ cells from 10-12 week old foetuses
Induced pluripotent stem cells (iPS cells)
- artificially created from somatic cells
Definitions of SC potency - flow
Zygote = totipotent
Blastocyst -> ES cells = pluripotent
Epiblast -> epiSC = pluripotent
Late embryo/early foetus (primordial GCs) -> EG cells = pluripotent
Adult stem cells = multipotent or unipotent
-> skin, CNS, bone marrow, other -> some stuff = iPS, pluripotent
Waddington’s ski slope model
add
Mesenchymal stem cells
multipotent
give rise to osteblasts, adipocytes and chondrocytes
Hematopoietic stem cells
multipotent
give rise to common lymphoid (» T/B-lymphocyte, NK cell) or myloid progenitor (» Monocyte/macrophage, granulocytes, platelets, and erythrocyte)
Niche for HSCs?
Endosteal surface of the bone
Functions of adult stem cells
Proper tissue organisation
Response to demands of growth/repair - some restrictions on developmental potential
Limits imposed by powerful molecular restraints on gene expression, heritable during many rounds of cell division
ASC may show relaxation of restrictions in altered environment - plasticity.
Blastocyst stage
Body plan not apparent
Many cells don’t form new human - give rise to placenta etc, support pregnancy
Embryo does not represent a single individual (can be twins, up to 14 days)
No precursors of nervous system yet
Not possible to predict if developed to term
Ethical considerations
dsfds
Definition of an Embryonic Stem Cell
Originates from a pluripotent cell population
Stably diploid, karyotypically normal in vitro
Can be cultured indefinitely in its primitive state
Can be differentiated into tissues of the three embryonic germ layers: endoderm, mesoderm and ectoderm in vitro or in teratomas
Can give rise to any cell in the body when injected into a host blastocyst (unethical and illegal with human ES)
Making chimeric mice with mouse ESC
1) ESC derived from pigmented strain of mice
2) Recipient blastocyst from albino strain of mice held by holding suction pipette
3) ESC in pipette injected into cavity of blastocyst
4) Injected cells become incorporated in inner cell mass of host blastocyst
5) Blastocyst develops in foster mother into a healthy chimeric mouse
Teratoma formation
Surrogate test for pluripotency
Xenografts in SCID mice
Ability of a human ES cell line to form teratomas is the best test of pluripotentiality
Implant cells for tsting under testis capsule of SCID mice or intramuscular
Monitor animals weekly, starting at around 4w weeks for further development
Remove tumours and fixed in formalin, send for routine histological processing
What can the human ES cells differentiate into?
Cartilage, cardiomyocytes, neurons, glandular epithelium, gut, blood, muscle and pigmented epithelium
Characterisation of human ESC
Biological properties - differentiation into derivatives of all three germ layers (teratoma, embryoid bodies)
Morphology and cell cycle
Immunological markers
Gene expression: transcripts for generic markers of pluripotent stem cells
GCTM-2, TG 343, TRA-1-60, TRA-1-81
Antibodies, targets:
Large surface keratan sulphate/chondroitin sulphate proteoglycan
OCT-3/4, Nanog, Sox2
Transcription factors, can induce expression of each other
Important for maintaining the self-renewing undifferentiated state of the inner cell mass in the blastocyst, as well as ESC
SSEA-3, SSEA-4
Cell surface glycolipids
Key players in cell signalling
Identifies many types of mammalian cells with pluripotent and stem cell-like characteristics.
TG30, CD9
Cell surface proteins
CD9: tetraspannin required for maintenance of pluripotent state in mouse ES cells`
RPE program
Retinal pigment epithelial cells from hESC
Easy to identify
Small dosage vs. other therapies
No risk of rejection at injection site
Ease of administration
May positively impacter > 200 retinal diseases
Pros and cons: Ebryonic ( and iPS) stem cells
Pros: unlimited expansion and differentiation, ethical issues - patient tissue derived (iPS)
Cons: Genetic instability, forms teratomas (tumours), ethical issues (embryo)
Pros and cons: Adult stem cells
Pros: Genetically stable?, no tumorigenesis?, patient tissue derived (BM), allogeneic possible, immunosuppressive
Cons: limited expansion, restricted differentiation
