M6 Stem cell therapy

Question 1

Transplantation types of cells

Answer 1

• Allogenic (heterologus) transplantations:
  
  • Involves diff donor/host (need immune suppressive therapy to prevent reject of transplant)
• Autologous transplantation:
  
  • Only 1 individual needed (host&donor)

Question 2

ESC

Answer 2

• Able to differentiate into any cells in the body (endo/meso/ecto)
• Umbilical cord cells → good for isolation of mesenchymal + haematopoietic SC

Question 3

Population and induction of SC

Answer 3

• Every tissue has a population of it’s own SC
• Inducible pluripotent SC → induce back of SC lineage

Question 4

What are the majority of SC trials based around?

Answer 4

• Majority of SC trials are based around hematopoietic SC

Question 5

Skeletal muscle SC (Adult SC population)

Answer 5

• Capable of self-renewal and differentiation into muscle
• B/w sarcolemma and basal lemma
  
  • Perfect position to receive signal from muscle and from external env
• Most in quiescent state (maintained in 2 to 3 years) → activated with injury/insult
• Co-localisation of satellite cells with capillaries
  
  • Satellite cells exist in aerobic state
• Satellite cells are key for muscle regeneration

Question 6

Transcriptional regulation of SC

Answer 6

• Quiescent cells express high levels of Pax7 → enter cell cycle and upregulate myoD
• MyoD → essential for specialising cell lineage → commited to become a muscle fibre

Question 7

Different pathway of quiescent cells

Answer 7

• Able to return to quiescent state to regenerate SC pool
  
  • Does not express myoD
• OR moves on to differentiation into myocytes → myotube → myofibre

Question 8

Transplanted cell survival

Answer 8

• 90-95% of cells die within the first 24hr
• HOWEVER, most cells survive if transplanted directly (Taken out freshly from donor and injection without culturing)
  
  • Shows problem with in vitro culturing of stem cells

Question 9

Cells which had poor survival were reliant on?

Answer 9

• Cells which relied heavily on glycolysis → poor survival
• Cells which did not rely on glycolysis → good survival

Question 10

Why do freshly transplanted cells have much better survival?

Answer 10

• By genetically preventing differentiation to myoD
• Able to transplant them wit 2-3x efficiency
• Way to culture cells in vivo and expand cells without it comitting to myogenic lineage → goal

Question 11

Glycolysis in SC proliferation

Answer 11

• Glycolysis is essential in proliferation
  
  • Generates the biomass for proliferation (building blocks - including nucleotides, phospholipids, AA)

Question 12

What do quiescent cells and proliferating cells rely on?

Answer 12

• Quiescent cells heavily relied on fatty-acid oxidation
• Proliferating cells heavily relied on glyocolysis

Question 13

Inhibiting myogenic specification in culture without compromising proliferation

Answer 13

1. Culturing cells in different mediums
2. TXNIP

Question 14

Culturing cells in different mediums

Answer 14

• High glucose (high rate of glycolysis)
• Low glucose (rate of glycolysis decrease)
• Galactose (rate of glycolysis decrease + increase oxidative phosphorylation)
  
  • Decrease expression in MyoD

Question 15

Result of decreasing the reliance on glycolysis pathway

Answer 15

• Low glycolysis groups increases proliferation and generation of biomass
• Able to reduce specification of cells to the myogenic lineage

Question 16

TXNIP

Answer 16

• ↑TXNIP expression → Inhibit glycolytic ability of cells → ↓commitment to myogenic lineage
• BUT overexpression of TXNIP → Reduces myogenic specification (decrease MyoD) BUT stop proliferating
  
  • Becomes senescence (irreversible G0 state)
• TXNIP is heavily expressed in quiescent cells
• No role involved for TXNIP in proliferation