Stem cells Flashcards
Progenitors
Cells that proliferate for a limited number of cycles before differentiation
Cell lineage
developmental history of a differentiated cell as traced back to the cell form which it arises
Blastomere
the cell type of the early embryo that is generated by cleavage of the zygote
Blastocyst
the spherical embryo at the time of implantation, consisting of the primary tissue types
What are the three primary tissue types?
trophectoderm
epiblast
primitive endoderm
Trophectoderm
precursor of the placenta
Epiblast
the founding tissue of the embryo proper that gives rise to all fetal tissues
Primitive endoderm
extraembyronic tissue that initially covers the epiblast surface and later gives rise to the yolk sac tissue
Potency
ability of a cell to differentiate into one or more cell types
Totipotency
ability of a cell to give rise to a fully functional organism, both embryonic and extraembryonic tissues
Pluripotency
ability of a cell to develop into all embryonic but not extraembryonic cell types, including the germ cells
What are the hallmarks of pluripotency?
expression of pluripotency TFs
teratocarcinoma formation
Germ layers
the first specialised precursors of different embryonic cell types
Ectoderm
Surface, neural and neural crest
makes skin, PNS and CNS
Mesoderm
axial, paraxial, intermediated and lateral
makes blood, heart, muscle and kidneys
Endoderm
gut and internal organs
makes liver, pancreas, intestines
How is cell lineage decided during gastrulation?
in a regionalised manner- so the location of a cell predicts its identity in response to distinct signals that activate lineage specific TFs
Epithelium
cells that line the surface of a structure, characterised by tight junctions and polarised morphology
Epithelial to mesenchymal transition
a process during which cells lose their epithelila characteristics, gain a less regular appearance and become migratory
breakdown of basement membrane and loosening of cell cell contacts
Neuromesodermal progenitors
bipotent- paraxial mesoderm
future skeletal muscle, bone, cartilage and vertebrae
OR spinal cord
What do defects in neuromesodermal progenitors lead to?
spina bifida
currarino syndrome
sacral agenesis
sponylocostal dysotosis
What do haematopoietic stem cells give rise to?
all blood cell types
Aorta-gonad-mesonephros region
embryonic tissue originating from the para-aortic sphlanchopleura and consisting of the dorsal aorta and urogenital ridges
involved in generating HSCs prior to onset of haematopoisis in fetal liver
Why are embyros and stem cells difficult to study?
they have small cell numbers and have difficulties with in utero development and ethics
How do you capture pluripotent embryonic stem cells?
plate on layer of feeder cells (irradiated stromal cells from later embryos)
once they have divided a few times disaggregate and replate
What are the critical signals to maintain cells in a self-renewing undifferentiated state?
mice- leukaemia inhibitory factor
humans- FGF2 and TGF beta
What are the main pluripotency factors expressed by ESCs?
Oct4
Nanog
Sox2
3D
remove signals that keep cells in an undifferentiated state such as BMP and LIF (mice) and FGF2 (humans)
grow in aggregates in the presence or absence of signals
Advantages of 3D approach
recapitulates more accurately ambryonic development
Disadvantages of 3D approach
difficult to observe or dissect the role of individual signals
2D / adherent
plate a defined number of cells on the right substrate or ECM
remove signals that keep cells in an undifferentiated state such as BMP/LIF/FGF2
grow in a defined medium with appropriate amounts of signals
Advantages of 2D approach
more tractable system for live imaging and is easier to test the role of specific signals
Disadvantages of 2D approach
loss of cell interactions that may occur in vivo
Microcephaly
neurodevelopmental disorder in which infants are born with an abnormally small brain due to various autosomal recessive mutations
causes neurological defects and seizures
Capturing cells in vitro
dissociate cells and plate on laminin in the presence of cytokines FGF2 and EGF
single cell can generate identical daughter cells and differentiate into glia/neurons after removal of cytokines
Differentiated cells characteristics
do not divide or do it less frequently
limited reversibility and plasticity
restricted capacity to cope with injuries, insults or wear and tear
Conrad Waddington’s epigenetic landscape
illustrates the progressive nature of differentiation
What organs in the body have regenerative capacity?
liver
epithelial cells
skeletal and cardiac muscle
brain blood cells
Adult stem cells
stem cells present in adult tissues and organs that retain the characteristics of stem cells such as self renewal and potency
but are usually unipotent or multipotent, limited cell fate decision
Cellular homeostasis
constant or periodic generation of new cells to replace old damaged or cying cells
or the addition of new cells as needed by regeneration/replacement by adult stem cells
Classical stem cell division
produces transit amplifying or progenitor cells which then differentiate
Stem cell assymetry
one progenitor cell is produced
Population asymmetry
entire cell population has some cells that will become progenitor cells and some that remain as stem cells
Stem cell niche
specialised specific microenvironment regulating adult stem cells physically and chemically
How does the stem niche regulate stem cells?
physical- cell adhesion and ECM
chemical- secreted proteins (paracrine, juxtacrine, endocrine, NTs)
Epigenetic regulation
histone modifications and methylation influencing the expression of stem cell genes
Transcriptional regulation
networks of TFs that regulate stem cell quiescence, proliferation, differentiation and self renewal
Cytoplasmic determinants
assymetric distribution of proteins that govern the mode of cell division
Drosophila creation of oocyte
ovarian germline stem cells divide to generate one stem cellm and one cytoblast cell
these divide into many more before selection of the oocyte
How does BMP signalling control germline stem cell differentiation?
cap cells produce TGF beta ligands
these activate BMP signalling
this represses BAM which is required for differentiation
What causes germline tumours?
overexpression of DPP or loss of BAM
What causes germline stem cell differentiation?
loss of dpp or overexpression of BAM
How do skeletal muscle cells repair muscle tears?
assymetric cell division in stem cell maintainance
Basal lamina
part of the stem cell niche, layer of ECM secreted by epithelial cells
remodelling of this supports assymetric cell division
How are adult stem cells anchored to their niche?
cadhenin mediated physical cell-cell adhesion
integrin mediated cell-ECM interactions
How do cadhenin mediated cell cell adhesions work?
a-catenin and b-catenin associate with intracellular domain of cadherins
help to cluster cadherin molecules and form adherens junctions
How do integrin mediated cell-ECM interactions work?
intracellular domains of integrins interact with actin cytoskeleton network through talin proteins to cluster integrin molecules together
What other properties do stem cell niches have?
signalling molecules are secreted by the cell in high concs close to the stem cell causing self renewal and regulates proliferation
grow cells being used to treat a wound
prevention of premature ageing
How is asymmetric cell division important for cell polarity?
the cell that is away from the niche will not have any signals anymore so will be on the path of differentiation
How does competition play a role in quality control of stem cells?
there is still some cadherin and integrin signalling but if even a little bit of this is lost it will induce differentiatoin
Stem cell ageing
old stem cells lose cadherin which means that they are lost from the niche, so are more susceptible to infections, becoming more clonal, and mutations
Gut crypt structure
tube of cells with stem cell like cells in a niche at the distal end, and differentiated cells at the proximal end
Gut crypt structure
tube of cells with stem cell like cells in a niche at the distal end, and differentiated cells at the proximal end
stem cells surrounded by paneth cells and basement membrane
Paneth cells
cells surrounding the stem cells at the bottom of the crypt that secrete many different peptides and small molecules that regulate the gut microbiome, and signals to Lgr5
Lgr5
Receptor that responds to Wnt signalling
How can mini guts be made ?
single stem cell populations can be isolated and amplified, then given growth factors and Lgr5+
LGR-EGFP transgenic mice
instead of the crypt looking organised, the adenomas mean that paneth cells and the lgr5 transgene is expressed all the way up the crypt, as they are activated and therefore not in the right place
How can pluripotency be used to make organoids?
reprogramming skin fibroblasts or using embryonic stem cells
taking stem cells from gut epithelium and sorted using FACs
FACS
fluorescent activated cell sorting
What is the role of Wnt signalling?
induces local proliferation and EphB expression
local cell expansion
Eph-Ephrin repulsive force which generates bud formation
How is stem cell reprogramming used as an experimental tool?
research for intestinal stem cells, intestinal differentiation and epithelial infections
How is stem cell reprogramming used as a diagnostic tool?
for cystic fibrosis, mutational analysis in CRC
drug absorption and metabolism
How is stem cell reprogramming used as a therapeutic tool?
potential regenerative therapy for microvillus inclusion disease, ulcerative collitis and post-endoscopic mucosal resection
