ANGIOGENESIS in TE engineering

Question 1

Importance of vascularisation?

Answer 1

• Most tissues in body contain vascular network
• happens as a result of hypoxia
  cells differ in their sensitivity to oxygen

Question 2

Why is vascularis. a key bottle neck?

Answer 2

Need vessel and nourishment from blood to enable survival of cells in TE constructs

• avoids graft necrosis
• aids in innervation
• generates thicker tissues
• improves graft functioning

Question 3

Examples of macro, micro vessels? Which mediate exchange of nutrients?

Answer 3

• Macrovessels = veins and arteries
• Microvessels - arterioles and venules.
  Exchange of nutrients mediated by capillaries

Question 4

Vessel formation - what are the 3 types?

Answer 4

• Vasculogenesis –> do novo formation from progenitor cells
• Angiogenesis –> new vessel formation via extension and remodelling of existing blood vessels
• Arteriogenesis –> maturation of blood vessels via increasing the lumen size

Question 5

Vasculogenesis - outline?

Answer 5

Hemangioblasts form from the mesoderm. This leads to tube formation via rearrangement of cells, which forms primary capillary plexus

Question 6

Angiogenesis - what drives it? Outline process?

Answer 6

Driven by hypoxia
Hypoxia causes production of pro-angiogenic factors (eg angiogenic vascular endothelial growth factor - VEGF). This signals to the ECs which secrete enzymes that degrade the basement membrane and proliferate to create vascular sprouts which grow towards hypoxic signal and form new lumen.

Question 7

How does angiogensis operate as a negative feedback loop?

Answer 7

Once blood flowing through lumen of new vessel, the vessel becomes less hypoxic. Factors that promote angiogenesis switch on transcription factors that inhibit process

Question 8

Importance of VEGF in angiogenesis

Answer 8

VEGF - vascular endothelial growth factor –> proliferation of ecs, promotes their migration and differentiation into new vessels

Question 9

Importance of HIF in angiogenesis

Answer 9

Transcription factor that stimutes pro angio factors eg VEGF/PDGRF/TGF-a/EGF

Question 10

Importance of PFGRF in angiogenesis

Answer 10

Platelet derived growth factor - recruits and stimulates proliferation of pericytes and VSMCs

Question 11

Importance of Angio poietn in angiogenesis

Answer 11

Regulation of EC survival, sprouting and pericyte recruitment

Question 12

Importance of Matrix metaloproteinases in angiogenesis?

Answer 12

Basal lamina degradation, remodelling of ECM

Question 13

Roles of VEGFA and B in angiogenesis?

Answer 13

Tyrosine kinases –> bind to VEGFR1 to induce vasculogenesis

Question 14

Roles of VEGC and D ?

Answer 14

Bind to VEGFR2 –> stimulates EC proliferation, differentiation and survival and tumour angiogenesis

Question 15

What stimulates arteriogenesis and outline process?

Answer 15

Vessels remodelled in response to fluid shear stress (signals to cells regarding blood flow)
Endothelial cells release growth factors which stimulates proliferation of ECs and prolif of SMCs, as well as matrix remodelling (small arterioles become larger by their lumen increasing in size)

Question 16

What are the main strategies for vascularisation in TE?

Answer 16

• scaffold design and functionalisation (strategy for facilitating vascular ingrowth)
• pre vascularisation strategies

Question 17

What would ideal scaffold for vascular ingrowth be? why?

Answer 17

Porous scaffolds created to promote vascular ingrowth. Allows functionalisation of scaffold!!!
Can then decorate the scaffold with growth factors that allow vascularisation ef VEGF/PDFG/bFGF. Can induce growth in a controlled manner

Question 18

Pre vascularisation strategies - how do they work?

Answer 18

TE construct is cultured in vivo to build a pre vasularised structure
Pre vascular network can recreate a connection with an existing blood vessel rather than having to undergo new angiogenesis. In this technique, scaffold is applied and endothelial cells spontaneously self-assemble into capillary-like structures.

Question 19

Disadvantages to in vivo (pre) vascularisation strategies?

Answer 19

• Sourcing of cells
• Scaffold must be implanted into patient at easily accessible, well vascularised tissue to allow ingrowth from host
• Microvessels are ingrown from the host
• After vascularisation, transferred to the defect site.
• Scaffold must then be removed. 3 SURGERIES IN TOTAL!!!!

Question 20

Flap technique - what is it and why may it be preferable?

Answer 20

Scaffold implanted into muscle flap
Microvessels ingrown from the host. Once this occurs, entire flap transferred to the site of repair
The vascular pedicle of the flap is surgically anastomosed to host vessels

Question 21

Advantages and key problems of flap technique?

Answer 21

• Large vessels can be surgically sutured together immediately to allow immediate blood flow
• Key problem is that may have to sacrifice a large amount of muscle tissue!

Question 22

AV LOOP TECHNIQUE

Answer 22

1) Uses a vein or synthetic graft to form a shunt loop between an artery and vein
2) AV loop leads to spontaneous sprouting of vessels
3) The loop is placed in a protected chamber
4) When vascularized it can be transferred to the repair site

Question 23

Advs and Disadvs of AV LOOP technique?

Answer 23

+ Allows vascularisation of the tissue construct
+ No major morbidity at donor site
+ Not embedded in surrounding tissue

• labour intensive as independent of vessel growth in the body
• multiple surgeries as the vein and artery must be removed from the patient

Question 24

Advantages and disadvantages of in vitro pre vascularisation techniques

Answer 24

\+ Doesn't rely on host ingrowth
\+ No extra surgeries 
- complex strategy and more labour intensive 
- strategy varies from tissue to tissue 
- not as fast

Question 25

Advs and Disadvs of in vivo pre vascularisation techniques?

Answer 25

+ involves direct perfusion of blood after microsurgery to join major vessels
+ mature and organised vasculature
- extra implantations and surgery often necessary
- need to find proper location for pre vascularisation
- scaffold may be filled with fibrous tissue during initial implantations