Lecture 2 - Transdifferentiation Flashcards
What is transdifferentiation?
Transdifferentiation is the conversion of one cell type to another, e.g. pancreas cell to liver cell
What is metaplasia?
Metaplasia is conversion of one cell type to another and can include the conversion of one tissue stem cell to another tissue stem cell, e.g. oesophageal to intestinal
What is the difference between transdifferentiation and metaplasia?
Transdifferentiaation specifically refers to conversion of one differentiated cell type to another without going through a less differentiated state, while metaplasia refers to any conversion of one cell type to another.
What’s Barrett’s oesophagus?
An acquired condition characterised by metaplastic replacement of the normal squamous cells by intestinal-type columnar cells.
Why does Barrett’s oesophagus occur?
Chronic gastroesophageal reflux disease (GERD)
Why is transdifferentiation important to study? (3 reasons)
- Transdifferentiation predisposes to neoplasia (cancer) - understanding molecular and cellular basis of metaplasia may help early diagnosis and imporve therapeutic targets
- Identifying transcription factors responsible for transdifferentiation may help in reprogramming stem cells toward differentiated cell types
- Transdifferentiation informs us about normal development of the two cell types that interconvert
GERD action
- Sphricter relaxed
- Acid and bile bile from stomach to oesphagus
- Disruption of epithelial cells and replacement
- Repeated exposure causes replacement to columnar epitheliam (all 5 different types seen)
Patient cancer statistics with Barrett’s metaplasia
40-100 fold increased chance of develping oseophageal cancer (with a 5 year survival ate of 10%)
What are morphogens?
Substancces that establish a graded distribution and elicit distinct cellular responses in a dose-dependent manner.
What is the function of morphogens?
Provide individual cells within an area with positional information, which is interpreted to give rise to spatial patterns.
Example of morphogens involved in transdifferentiation
Neural context:
- BMP inhibition and FGF signaling in neural progenitors activate proneural genes (master TFs) like neurogenin and NeuroD
- NeuroD converts non-neuronal cells (like pancreatic cells or astrocytes) into neurons
Muscle context:
- Wnt and Sonic HedgeHog signaling promotes MyoD expression in the somites
- MyoD is master regulator of skeletal muscle differentiation and converts fibroblasts into muscle cells
Criteria for confirming transdifferentiation
- Describe the phenotype before and after switch (loss of one phenotype and gain of another) - morphology, sequencing, protein expression, gene expression, drug testing
- Demonstrate lineage relationship between ancestor (starting cell) and ddescendant (finishing cell)
How to analyse cell phenotype?
- Morphology
- Gene expression (molecular level)
- Protein level (immunostaining or WB)
- Functional characterisation
How can linegae analysis be completed?
‘Label’ cells with a reporter (e.g. GFP), exclude any stem cell intermediates using stem cell markers
Key principles:
1. Heritable markers - markers must be passed onto progeny
2. Specificity - markers should not diffuse or be transferred to neighbouring cells
3. Stability - should allow analysis of linegaes over extended period of time
4. Non-toxicity - should not interfer with accuracy of lineage analysis
Symptons of liver disease
- Oedema (refuction in albumen production)
- Capid medusa (blood vessels opened due to portal tension)
- Jaundis (high bilirubin levels)
- Matastrolis/high oestrogen
- Erectile dysfunction
- Testicular atrophy
Example of model for induction of hepatocytes in pancreas (copper depletion-repletion)
Rao et al (1986) in rat
Example of in vitro model for transdifferentiation of pancreas to liver
Rat pancreatic AR42J-B13 cells
What are AR42J-B13 (B13) cells?
Pancreatic cells derived from azaserine-treated rates
What is the function of AR42J-B13 (B13) cells?
Express exocrine and neuroendocrinne properties, e.g. amylase, synaptophysin and neurofilament
Amylase expression is increased when cultured with synthetic glucocorticoid dexamethasone (Dex)
Can induce the formation of insulin-secreting B-cells with activin and hepatocyte growth factor (HGF)
May represent pancreatic progenitor cell
AR42J-B13 cell phenotype when treated with Dex
AR42J-B13 cells express a pancreatic phenotype under control conditions and a hepatic phenotype following treatment with Dex
What does Dex stand for?
Synthetic glucocorticoid dexamethasone
Features of hepatic cell
- Amylase expression decreased
- Can be binucleic centrally located
- Polygonal
What do transdifferentiated hepatocytes express?
Many proteins found in normal cells, e.g. phase I (CypA1) and Phase II (UGT) drug metabolising enzymes
Examples of gene expression of hepatocytes induced from pancreatic cells
Fibrogenin family = a-fibrinogen - 278, b-fibrinogen = 336
Phase I metabolism = Cyp3A1 - 353
Gluconeogenic enzymes = Glucose-6-phosphatase - 28, pyruvate carboxylase = 28
