Cartilage Flashcards
1
Q
How does cartilage change at different depths?
A
- material properties change as determined by biochemical nature and ECM content + organisation
2
Q
What is the ECM made up of?
A
COLLAGEN
PROTEOGLYCANS
OTHER MATRIX PROPERTIES = FN, link protein, bioglycan, decorin, hyalluronan
3
Q
What are the types of collagen?
A
2,5,6,9,10,11
4
Q
What are the types of proteoglycan
A
Aggregan
Versican
Lubrican
5
Q
What are the roles of the ECM?
A
- maintain mechanical properties of tissue
- sequesters growth factors and proteinases to specific compartments
- interacts with chondrocytes which regulate cell activity
6
Q
How does ECM maintain mechanical properties of tissue?
A
- type 2 collagen helps withstand tensile + shear forces
- proteoglycan = solute flow + tissue deformation
- water
7
Q
Define territorial and interterritorial
A
Territorial = matrix closest to the chondrocyte Interterritorial = matrix between chondrocytes
8
Q
What is the diameter of chondrocytes?
A
10 um (micrometers)
9
Q
What is mechanical loading?
A
- biochemical and mechanical signals
- regulates ECM production and/or breakdown
- maintains health of tissue
10
Q
Anabolic cell molecules?
A
- GF
- anti-inflammatory cytokines
- TIMPs
11
Q
Catabolic cell molecules?
A
- cytokines
- MMPs
- signaling mediators
12
Q
What type of articular cartilage injuries occur in young and active population?
A
- sporting and work related
- road traffic accidents
13
Q
Non pharmalogical therapies for OA
A
FOR MILD SYMPTOMS
- patient education
- physical/occupational therapy
- exercise and lifestyle
14
Q
Therapeutic approached therapies for OA
A
- NSAIDs
- Rofecoxib
- celecoxib (Pfizer)
- Lumiracoxib
15
Q
Disease modifying agents for OA
A
- nutraceuticals (glucosamine)
- DMOADs (cytokines, MMPs)
- licofelone, COX
16
Q
How to use autologous cell implantation?
A
- health cartilage
- isolate cells and expand
- inject cells under periosteal graft
17
Q
How to do cartilage repair?
A
- remove biopsy
- isolate chondrocytes
- expand in monolayer culture
- bank cells for future use
- seed in scaffold
- biochemical factors (in vitro formation of neo-tissue)
- implant functional device
18
Q
Define mechanotransduction
A
activation of cell signalling
apply mechanical load to improve response of system/alter pathways and cell response
19
Q
Physical effects at different levels
A
TISSUE -> interstitial fluid flow, hydrostatic pressure, osmotic, pH
CELLULAR = volume, deformation
INTRACELLULAR = nucleus + cytoskeleton deformation, second messemger pathways, signalling
MOLECULER = gene, protein expression
20
Q
Types of loading configurations?
A
- tension
- compression
- shear (fluid or mechanical)
- hydrostatic
21
Q
Advantages of chondrocytes agarose models
A
- reproducible
- well established
- cells adopt spherical morphology
- maintain chondrocyte phenotype
- mechanically characterized system
- permits application of physiological levels of cell strain to chondrocytes
- enables examination of effects of cell deformation on signaling pathways with and w/o multiple factors
- permits ECM component synthesis
22
Q
Disadvantages of chondrocyte/agarose model
A
- interactions of matrix network are lose
- could lead to altered mechanosignalling events
23
Q
Advantages of the flexcell system
A
- enables examination of cell stretch on complicated signalling events involving TF, ion channgels, mRNA, receptors etc.
24
Q
Disadvantages of flexcell system
A
- physiological relevance of stretch magnitude
- uniformity of stretch and other substrate
- loss of chondrocyte phenotype
- loss of response to mechanical/biochemical signals
25
Advanatges of compressive explant system
- interactions of the matrix network are maintained in explants
26
Disadvantages of compressive explant system
- not reproducible model system = heterogenity of tissue means difficult to control cellular strain, different local stresses and strains percieved by chondrocytes, so biochemical response will vary
- difficult to control geometry of full thickness tissue following join excision
27
Examples of abnormal mechanical loading
- altered joint loading
- obesity
- bone remodelling
- trauma
28
Examples of abnormal physiology
- inflammation
| - immune response
29
Result of abnormal mechanical loading and abnormal physiology
- matrix damage
- abberant repair responses
- mechanical failure
- joint distraction, pain, disability
30
In vivo events causing initiation of OA to early OA
- mechanical injury
- hereditary factors
- ageing
31
In vivo events causing early OA progression to late OA
- inflammation
- repetitive injury
- subchondral bone changes
32
What type of tissue organisation is cartilage?
Hetergenous
- hyaline
- chondrocytes
- lacunae
- matrix
- collagen
- proteoglycans
33
What is the purpose of mechanical loading?
- essential for normal metabolism
- regulates ECM production/breakdown
- maintains tissue health
- biochemical and mechanical signals
34
How can cartilage be repaired?
Total knee replacement
35
What are some hypotheses on cell mechanotransduction?
- dynamic compression stimulates cell proliferation and proteoglycan synthesis in a frequency dependent manner
- response of chondrocyte/agarose constructs to dynamic compression depends on cell sub-population
- chondrocyte metabolism is dependent on length/type of compressive regime applied
- compression induced changes in chondrocytes/agarose constructs are enhanced by TGFB and are integrin mediated
- dynamic compression inhibits production of NO in the presence of IL-1B
- dynamic compression inhibits iNOS and COX-2 expression via distinct pathways in the presence of IL-1B
36
How can we test the effect of dynamic compression?
- full depth chondrocytes seeded in agarose and equilibrate in culture
- subject chondrocyte/agarose constructs to static strain (15%) and dynamic compression (15%) at different frequencies
- look at cell proliferation and proteoglycan synthesis
- use compressive bioreactor system
37
What is the effect of dynamic compression in chondrocyte/agarose constructs?
- dependent on type of loading (static or dynamic)
- static inhibits cell proliferation and proteoglycan synthesis
- dynamic stimulates cell proliferation and proteoglycan synthesis
- proteoglycan synthesis was dependent on type of frequency
- cell proliferation was frequency independent
38
How to test if response of chondrocyte/agarose constructs to dynamic compression is dependent on the cell sub-population?
- isolate superficial cells
- isolate deep zone cells
- seed separately into agarose gel
- apply static (15%) and dynamic (15%) strains
- frequency at 1Hz
- look at cell proliferation and GAG synthesis
39
Is the response of chondrocyte/agarose constructs to dynamic compressiondependent on the cell sub-population?
- yes
- superficial cells showed greater cell proliferation (mediated by NO and calcium)
- deep cells showed greater proteoglycan synthesis (mediated by fluid flow)
40
What are the different dynamic compression regimes and types?
```
REGIMES
- continuous
- intermittent
TYPES
- unstrained
- strained
```
41
What is the effect of continuous or intermittent compression on cell metabolism?
- large numbers of duty cycles stimulates proteoglycan synthesis
- shortest duration of intermittent compression maximally enhance cell proliferation
- uncoupled nature of metabolic response suggest that mechanical conditioning regimes can be fine tuned with compressive bioreactor to selectively stimulate key metabolic parameters
- temporal mechanical conditioning response is therefore important in engineering cartilage tissue
42
What are the principal receptors which percieve mechanical signals and activate signalling mediators?
- integrins (a5B1)
43
Which signalling mediators are involved in mechanotransduction process?
- calcium ions
| - NO
44
How to test effect of TGFB on dynamic compression?
- human cartilage extracted from patients undergoing ACI
- isolate chondrocytes, expand in monolayer and seed in agarose gel
- constructs subjected to dynamic compression (15%, 1Hz) for 48 hours under following conditions = untreated, TGFB, TGFB + GRADSP, TGFB + GRGDSP
45
What is GRGDSP?
competitive ligand for the a5B1 integrin
46
What is the effect of TGFB on dynamic compression?
- in presence of TGFB, dynamic compression enhanced proteoglycan synthesis and cell proliferation and inhibited nitrite release
- role of integrins as a mechanoreceptor in chondrocyte mechanotransduction has been demonstrated using the RGD peptide (GRGDSP)
- peptide acts as a competitive ligand for a5B1 integrin and so blocks response of chondrocytes to mechanical loading
- as a consequence compression induced stimulation of cell proliferation and proteoglycan synthesis was inhibited in the presence of the peptide
47
How does IL-1B effect dynamic compression?
- IL-1B stimulates NO release and inhibits cell proliferation and proteoglycan synthesis in chondrocyte/agarose constructs
- catabolic effects could be reversed with application of dynamic compression
- dynamic compression may be important in the modulation of IL-1B induced effects in articular chondrocytes
48
How does dynamic compression affect iNOS and COX-2 expression and production of NO and PGE2?
Our studies demonstrate an inhibition of iNOS and COX-2 expression and production of .NO and PGE2 release by dynamic compression.
