1 - TD Hepatocytes to Pancreatic cells Flashcards
Why is the ability to reprogramme hepatocytes to become pancreatic cells so useful?
- Liver = only organ in the body that regenerates its tissue
- So can harvest autologous hepatocytes, TD into pancreatic cells then transplant back
- Persons liver will grow back AND they have healthy pancreatic tissue = win win situation
What are the 2 functions of the pancreas?
- Endocrine - hormones
- Exocrine - digestive enzymes
What are the major symptoms of liver failure and what are they due to?
- Jaundice - RBC degradation by pancreatic enzymes is faulty so bilirubin builds up (by product) and it yellow
- Odema in abdomen - faulty liver means theres a loss of albumin and causes water to be kept in the blood resulting in odema
- Vissible blood vessel networks - odema pressure pushes the vessels to the skin surface
- Development of breasts in males (Gynecomastia) and testicular atrophy - abnormal sex hormones aren’t removed
What makes hepatocytes so preferable for TD to pancreatic cells?
- Liver and islet cell early development is identical until the pancreatic bug emerges
- Liver already expresses various TFs linked to pancreatic development
- Liver has a strong regenerative capacity, making them a scalable cell source
Give me 4 pancreatic TFs (pTFs) shown to reprogramme hepatocytes into IPCS
- PDX1
- NeuroD1
- Mafa
- TGIF2
At what stage in H-P TD is TGIF2 required and what does it facilitate?
Very beginning
Supresses hepatic identity
Induces pancreatic projenitor-like phenotypes
(unlocks the hepatic fate, allowing TD to IPCS to ocur)
At what stage in H-P TD is PDX1 required and what does it facilitate?
Early development
Participates in development, differentiation, maturation and proliferation of islet cells
At what stage in H-P TD is NeuroD1 required and what does it facilitate?
Middle development
Controls transformation into pancreatic endocrine precursor cells
At what stage in H-P TD is Mafa required and what does it facilitate?
Late development
Induces further maturation of endocrine precursors into islet cells
For in vitro TD, what are the advantages of using in vitro transcribed mRNA (IVT) over vector TF for delivery if TF’s for TD?
- RNA doesn’t have to reach the nucleus
- Doesn’t integrate with the genome, therefore no risk of insertion mutagenesis
- Protein expression can be accurately controlled by adding them @ specific times and dosages
What is the downside to using autologous cells for TD in treating T1D and is there any ways of getting around this?
T1D is an autoimmune disease
Therefore the body would still attack TD hepatocytes, meaning they still need immunosuppressants
BUT future advancements mean we’re now able to engineer the cells in a way that means they’re masked from the immune system
What are the downsides for using vectors to deliver TF’s to cells for TD?
- Can’t modulate or control the level of TF expression
- Cells have the potential bio-safety hazards due to insertion mutagenesis
What was the methods used for applying the IVT TF’s for H-P TD by Ma et al., in 2020?
Infected hepatocytes with the pTF’s sequentially from day 1-4: (3pTF group lacked TGIF2 so started from d2)
1. TGIF2
2. PDX1
3. NeuroD1
4. Mafa
Harvested the cells at day 6 to check for pancreatic-like signs
What was the effect of using TGIF2 + the 3 pTF’s?
- Cells treated with TGIF2 expressed pancreatic markers at a significantly higher amount Vs 3pTFs only
- Higher insulin response in changes in glucose concentration
- Only ones with TGIF2 were able to form islet-like cluster morphology
What was the conclusion surrounding the role of TGIF2 in H-P TD?
It’s the key unlocking H cell fate and allowing TD to occur
Overexpression in H results in transcriptional remodelling
This supresses the hepatic identity to unlock the potential for pancreatic projenitor-like phenotypes to be induces by pTFs
