Modelling tissue repair Flashcards
What are 2 extreme examples of wound healing?
Diabetic ulcer
Burns
What happens when burns heal?
Healing replaces matrix
Force that cells generate to replace matrix results in contracture
Affects function of tissue
Healing is of lower quality
What are the 4 stages of healing?
Inflammation
Re-epithelization
Granulation
Fibroplasia
What causes inflammation?
Cells rupture
Immune response leaves blood
What is re-epithelization?
Epithelia brings the wound closer together and seals it to protect host from extracellular environment
What is granulation?
Cells come in and replace hole produced by injury
Clot = first granulation tissue that traps cells to pevent from bleeing
Cells comin in proliferate, migrate and pull wound edges closer together by the production of ECM
Factors attract blood vessels to come in
Neural input comes along to bring sensation to the skin
What is fibroplasia?
Maturation of the healing wound beneath the surface of the skin
What are the 5 types of assays used to model tissue repair?
Scratch assay
Substrate wrinkling assay
Stress relaxed collagen gel assay
Free floating collagen gel assay
Culture force monitor
Boyden chamber
What are characteristic features of scratch assay?
Simple and commonly used
What are the steps to performin a scratchh assay?
Take cells from the body
Cells until they reach confluence
Scratch across the culture
Observe how the cells migrate to pull the gap together
How can the scratch assay be used in different ways?
Can add inhibitors and cytokines to look at their effect on wound healing
What are the disadvantages of scratch assay
Main regulators of wound healing are found underneath the skin
2D nature of the asssay is problemtatic -> can’t grow fibroblasts as they convert to collagen type I
What is substrate wrinkling assay?
When cells attach to the matrix and pull it together, they wrincle the ECM substrate
Cells accumulate forces whilst pulling the wound together
How is substrate wrinkling assay linked to diabetic ulcers?
Diabetic ulcers develop since the substrate being pulled by cells to heal wound is too soft
Excess release of MMPs made substrate too soft
How can we use substrate wrikling assay?
Can look at the amount of force generated by the cells via lasers
Map filament and actin concentration to the amount of substrate wrinkled
How is a stress relaxed collagen assay performed?
Let the fibroblasts attach to the matrix
After a period of time release the gel from the base
It will contract massively within minutes
Contraction depends on level of stress accumulated by the fibroblasts
How can a Boyden chamber be used to model wound repair?
See how cells migrate from one part of the set to another
Steps to perform a Boyden chamber assay?
Make chamber with cells on top
Substance at bottom of chamber
Observe how the cells migrate from the top to the bottom
What is Boyden chamber assay used for?
Effect of cytokines on cell migration
What is a floating collagen gel assay?
Cells are embedded within collagen wells
Floating on multi-well plates
When gel sets, cells attach and contract the cell matrix
Decreases the size of the matrix
What is a floating gel assay used for?
Gives an idea of how cells
Attach, contract and remodel a tissue
Look at the effects of cytokine on wound closure
What is a culture force monitor?
Collagen gel is placed in a rectangular form
One side is attached to a force transducer
One side is attached to a fixed point
How does the force transucer measure the tension in the fibroblasts?
Cells attach to the transducer and generate force
Force transducer is the only thing that can move
Causes displacement of the bar
Can correlate the displacement to the force produced by cells
What is the culture force monitor used to look for?
Looks at the effect of different cells on contracting the wound
