Lecture 8 - Healing 2

Question 1

What is the structure of proteoglycans?

Name an important proteoglycan

Answer 1

Core protein + glycosaminoglycans

Draw this

• Many -ve charges

Aggrecan

Question 2

What is the function of cadherins?

Answer 2

Adherence of two neighbouring cells of the same type

Question 3

What happens when cells aren’t bound by integrins?

Answer 3

Apoptosis is triggered

Question 4

Describe the function of proteoglycans

Answer 4

• Hold a lot of water, forming a gel-like structure
• Reservoir for GFs (bind GFs on their G3 region)
→ Modulate cell growth and differentiation

Question 5

What are the essential processes in healing?

Describe briefly which factors mediate these processes

Answer 5

1. Angiogenesis
   • EPCs: endothelial precursor cells
   • VEGF
2. Fibroplasia
   • Fibroblasts
3. Remodelling

Question 6

Describe angiogenesis

Answer 6

‘Explosion’ of new blood vessels

– Proliferation from existing vasculature –

1. VEGF binds to VEGFR on endothelium
2. Endothelial cells re-enter cell cycle
3. Sprouting of capillaries
4. Reorganisation, maturation

– Proliferation from EPCs from BM –

1. VEGF released from ECM stimulated EPCs in BM
2. EPCs migrate to vasculature
3. EPCs start to proliferate
4. New branches made
5. Stabilisation of vessels:
   • Pericytes
   • Smooth muscle
   • Connective tissue

Question 7

What is vasculogenesis?

Describe the process

Answer 7

Specialised form of angiogenesis during embryogenesis

Laid down for the first time

Question 8

What is fibroplasia?

Compare this with fibrosis

Answer 8

Formation of new connective tissue

Fibrosis is any abnormal laying down of connective tissue

Question 9

What is remodelling?

Answer 9

Maturation and reorganisation of fibrous tissue

Vascular granulation tissue → avascular scar

Question 10

Which stem cells give rise to new vasculature?

Answer 10

EPCs

Endothelial precursor cells

Question 11

Where does VEGF come from?

When is it induced?

Answer 11

ECM cells
Indued by:
 • Hypoxia
 • TGF-β
 • PDGF

Question 12

How is angiogenesis controlled?

Answer 12

Interactions between:
 • GFs
 • Vascular cells
 • ECM
Stabilisation of vessels

Question 13

Describe stabilisation of new vasculature and how this is initiated

Answer 13

Must be stabilised
• Pericytes
• ECM
• Smooth muscle

Initiated by:
• Angiopoietin
• PDGF
• TGF-β

Question 14

Which cytokine plays a central role in fibroplasia?

Answer 14

TGF-β

Question 15

What is the role of macrophages in fibroplasia?

Answer 15

• Clear debris

* Secrete GFs

Question 16

What happens to vasculature as the scar matures?

Answer 16

Vascular regression

Question 17

Compare healing of small and large wounds

Answer 17

Small:
1. Scab formation, neutrophil infiltrate
2. Regeneration of epithelium
NB Minimal inflammation

Large
1. Big scab
2. Fibrosis, replacement of tissue
3. Scar formation
NB Inflammation

Question 18

What is the definition of ECM?

Answer 18

• Macromolecules outside cells
• Formed by local secretion
• Assembled into network surrounding cells

Question 19

What are the general functions of the ECM?

Answer 19

• Reservoir for GFs
• Reservoir for water
• Scaffold

Question 20

What is the basement membrane?

Describe its structure

Answer 20

Specialised part of the ECM

Structure:
• Laminin
• Proteoglycan
• Collagen type IV

Question 21

What are the two groups of matrix components?

Answer 21

• Basement membrane

* Interstitial matrix

Question 22

What are the three types of molecules found in ECM?

Answer 22

• Fibrous proteins
• Adhesive proteins
• Gelatinous-like molecules

Question 23

What are some examples of fibrous proteins?

Answer 23

• Collagen

* Elastin

Question 24

What are some examples of adhesion proteins?

Answer 24

• Fibronectin

* Laminin

Question 25

What are some examples of gelatinous-like molecules?

Answer 25

* Proteoglycan | * Hyaluronan

Question 26

Why is collagen important in terms of abundance?

Answer 26

Most common protein in the animal world

Question 27

Describe the structure of collagen

Answer 27

* 3 right handed triple alpha helices | * Gly-X-Y repeating units

Question 28

How many types of collagen are there? | What are the two main groups?

Answer 28

27 types • Fibrillar • Amorphous

Question 29

What are some disorders caused by mutation in collagen?

Answer 29

Ehlers-Danlos syndrome Achondrogenesis Osteogenesis imperfect Alport syndrome

Question 30

Describe the formation of collagen

Answer 30

1. Protocollagen 2. Cleavage of ends to form Tropocollagen 3. Association of tropocollagen into collagen fibrils 4. Cross linking of collagen fibrils to form collagen fibres

Question 31

Describe the structure of elastic fibres

Answer 31

Elastin core | Fibrillin surrounding

Question 32

In which tissues are elastic fibres important?

Answer 32

Blood vessels Skin Uterus Lungs

Question 33

What is an important disorder resulting from mutation in fibrillin? List the features

Answer 33

Marfan Syndrome Features: • Very long bones • defects in organs where elastic fibres are important (heart valves, lungs, eyes, aorta) • Chronic inflammation

Question 34

Why is chronic inflammation seen in Marfan syndrome?

Answer 34

Fibrillin (in which there is a defect) is needed for regulation of TGF-B-dependent inflammation

Question 35

Where are the adhesive proteins found?

Answer 35

* Membrane bound (thus, receptors) | * Cytoplasm

Question 36

What are the four major families of adhesion proteins?

Answer 36

* ICAMs * Cadherins * Integrins * Selectins

Question 37

What are ICAMs?

Answer 37

Integral cell adhesion molecules

Question 38

Describe the structure of fibronectin

Answer 38

Two chains linked by disulphide bonds

Question 39

What are the functions of fibronectin?

Answer 39

* Cell migration | * Cell attachment

Question 40

What are some features of laminin? What is its structure? Where is it found?

Answer 40

* Adhesion molecule * Most abundant protein in the basement membrane * 3 chains, cross shaped Location: • Basement membrane

Question 41

What are the functions of laminin?

Answer 41

• Cell - ECM attachment Important for: • Growth • Survival • Motility

Question 42

Describe the structure of integrins

Answer 42

Two chains: • α • β

Question 43

Describe the function of integrins

Answer 43

``` Attachment: • Cell-cell • Cell-ECM Important for: • Development • Leukocyte extravasation ```

Question 44

How does fibronectin connect to cells?

Answer 44

Via Integrin

Question 45

Describe the structural interaction of integrins and cells Why is this structure important?

Answer 45

* Transmembrane integrin * Focal adhesion inside the cell * Actin skeleton associated with focal adhesion This structure has a role in signalling from outside the cell to the nucleus

Question 46

In which cellular junctions are cadherins found?

Answer 46

Desmosomes

Question 47

Describe the structure of hyaluronan | Draw this

Answer 47

Long protein core with proteoglycan associated. | Slide 25

Question 48

What gives a tissue its turgor pressure? | What does this mean?

Answer 48

Hyaluronan | Able to resist compressive forces

Question 49

What are the general functions of VEGF?

Answer 49

* Angiogenesis * Vascular permeability * Endothelial proliferation and migration

Question 50

When does fibrosis normally occur?

Answer 50

In chronic disease

Question 51

What is new granulation tissue?

Answer 51

This is new connective tissue

Question 52

What is seen in granulation tissue?

Answer 52

* Tiny blood vessels | * ECM

Question 53

Which cells are the main players in fibroplasia?

Answer 53

Fibroblasts

Question 54

Why is TGF-β important for fibroplasia?

Answer 54

* Recruitment & proliferation of fibroblasts | * Increased synthesis

Question 55

Which molecule do fibroblasts lay down in fibroplasia?

Answer 55

Fibrillar collagen

Question 56

What does the outcome of healing depend on?

Answer 56

* Severity of injury * Duration of injurious stimulus * Tissue type * Patient factors (immunosupression etc.)

Question 57

What is the process that predominates in small wound healing? Compare this with large wound healing

Answer 57

Small wound: epithelial regeneration • minimal inflammation Large wound: fibrosis • chronic inflammation occurs

Question 58

What do the chronic inflammation cytokines lead to?

Answer 58

Collagen synthesis

Question 59

What are the chronic inflammation cytokines?

Answer 59

TNF IL-1 IL-4 IL-13

Question 60

What does decreased metalloprotinase activity lead to?

Answer 60

Decreased collagen degradation

Question 61

What are the steps in generic wound healing?

Answer 61

1. Demolition (injurious stimulus) 2. Proliferation 3. Migration 4. Synthesis 5. Remodelling

Question 62

List some of the components of the interstitial matrix

Answer 62

* Fibrillar collagens * Elastin * Fibronectin * Proteoglycans

Question 63

Compare location of fibronectin and laminin

Answer 63

Laminin: basement membrane Fibronectin: interstitial matrix (stromal)

Question 64

What is the stroma?

Answer 64

AKA the interstitial matrix of the ECM

Question 65

Which GF and receptor is most important in angiogenesis? | What does it bring about

Answer 65

VEGF: ligand, released by ECM VEGFR-2: receptor, located on endothelial cells Brings about proliferation of endothelial cells

Question 66

Why is the ECM important in angiogenesis? | What are the important components of the ECM?

Answer 66

``` Direction of migration of endothelial cells Components: • Integrins • Matricellular proteins • Proteases ```

Question 67

Describe the pathway to fibrosis

Answer 67

1. Chronic inflammation 2. Chronic activation of macrophages and lymphocytes, which produce: 3a. GFs 3b. Cytokines (TGF-beta) 3c. Decreased MMPs 4a. GFs stimulate fibroblasts 4b. Cytokines stimulate fibrogenic cells to make collagen 4c. Decreased MMPs means decreased collagen destruction 5. This all leads to collagen deposition, and thus fibrosis