Stem cells

• A stem cell is a cell that isn't differentiated; it can divide by mitosis to produce more stem cells or differentiate into specialised cells
• Self-renewal is the ability of stem cells to divide and produce more stem cells of the same type as the original mother cell
• Quiescence is a reversible exit from the cell cycle
• A stem cell niche is the microenvironment of a stem cell that provides signals needed for its survival and differentiation. The cell types surrounding the stem cell dictate the cell types the stem cell can differentiate into
• A cell lineage is the history of a cell from its origin to its final fate
• Stem cell potency is the ability of a stem cell to differentiate into other cell types. A highly potent stem cell is one that can differentiate into a wide range of cell types
• A pluripotent stem cell can differentiate into all the cells of the embryo except extraembryonic tissues.
• A multipotent stem cell can differentiate into a few cell types specific to the tissue it is localised in
• Pluripotent stem cells are identifiable as they contain all three germ layers: ectoderm, endoderm and mesoderm
• Cell plasticity is the ability of a cell to change its shape and function (phenotype) in response to environmental cues
• Positional differentiation refers to when a cell's position in the body directs its differentiation
• Cellular senescence is a triggered, stable cell cycle arrest.
• Progenitor/precursor cells are immature cells that haven't yet fully differentiated
• Transit amplifying cells are undifferentiated cells following a stem cell division
• The main ethical concern regarding stem cell research is the destruction of preimplantation embryos
• The four main transcription factors that can transform fibroblasts into induced pluripotent stem cells are:

  • Oct3/4
  • Sox2
  • c-Myc
  • Klf4
• Researchers want to induce pluripotency without transcription factors due to their oncogenic nature and cancer risk
• Pluripotent stem cells are extensively screened to ensure purity and quality
• Immunocytochemistry is used to visualise proteins using antibodies that are labelled with a fluorescent or enzymatic tag
• Immunocytochemistry is used to identify pluripotent stem cells by binding antibodies to marker proteins e.g. transcription factors Oct4, Sox2, and other proteins such as SSEA1 and NANOG. The antibodies are specific to the marker proteins and are labelled for identification e.g. with a fluorophore
• Reverse transcription PCR is used to detect presence of a marker protein to detect pluripotency. RNA sample -> cDNA gets amplified. This quantifies the RNA to prove that the marker protein is being transcribed
• Gapdh is used as a housekeeper gene when assaying protein expression
• Transcriptomic analysis allows scientists to quickly identify groups of genes expressed in stem cells. It is a good first step for further investigation to detect stem cell marker proteins.
• Bisulfite genome sequencing indicates where genes are repressed - identifies where marker proteins necessary for inducing pluripotency are not present.
• iPSCs are also detectable by the teratoma test, as teratoma formation requires all three germ layers; these are only present in pluripotent stem cells
• The teratoma test works like this:

  • Nude mice (lacking an immune system) are injected with pluripotent stem cells
  • This should lead to development of teratoma
• A neural stem cell is a cell that can differentiate into any type of neuron.
• In early embryonic development, cells commit to either the ectodermal lineage to form skin cells or the neural lineage to form the central nervous system
• During embryonic development, there are high levels of mitosis and no cells are at G0. These cells are multipotent stem cells; there is no specialisation.
• Later in development, regionalisation takes place; the embryo is divided into different specialised groups, controlled by morphogens
• Morphogens are concentration dependent factors that control the cell lineage of multipotent stem cells during development - i.e. dictates what cell types the stem cells will differentiate into
• Patches of specific cadherins are often expressed during embryonic development as they cause cells to stick together to influence them to differentiate into the same cell type
• Overproduction of neural progenitor cells allows those that develop in the wrong places to be destroyed. This ensures humans start off life with a robust CNS
• CNS development begins once the neural tube closes. This happens as epithelial cells receive a signal to become the neural plate border; these cells display specific clathrins which interact with cells on the other neural plate border. The neural plate borders interact and produce the neural tube
• Epigenetic switches cause neural precursor cells to switch to glial precursor cells.
• Neural cell synthesis is known as neurogenesis
• Glial cell synthesis is known as astrogliogenesis
• To study neurodevelopment in vivo (including the switch from neural to glial cells), organoids can be developed. Organoids are derived from stem cells in vitro and produce three-dimensional parts of tissues. This allows the human cerebral cortex to be modelled in vitro.
• Organoid development:

  • Stem cells are cultured in suitable medium – e.g. DMEM, L-glutamine, antibiotic (from practical).
  • Addition of signalling proteins to induce the culture to differentiate into desired stem cell types.
  • Testing to check successful differentiation.
  • Culture is grown into 3D desired shape – e.g. by embedding cells in a hydrogel matrix.
• Patterned expression of transcription factors enables regions of the brain to develop, responsible for different functions.