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Tissue Engineering & Regenerative Medicine > Cartilage > Flashcards

Cartilage Flashcards

1
Q

How does cartilage change at different depths?

A
  • material properties change as determined by biochemical nature and ECM content + organisation
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2
Q

What is the ECM made up of?

A

COLLAGEN
PROTEOGLYCANS
OTHER MATRIX PROPERTIES = FN, link protein, bioglycan, decorin, hyalluronan

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3
Q

What are the types of collagen?

A

2,5,6,9,10,11

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4
Q

What are the types of proteoglycan

A

Aggregan
Versican
Lubrican

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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
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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
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7
Q

Define territorial and interterritorial

A 
Territorial = matrix closest to the chondrocyte
Interterritorial = matrix between chondrocytes
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8
Q

What is the diameter of chondrocytes?

A

10 um (micrometers)

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9
Q

What is mechanical loading?

A
  • biochemical and mechanical signals
  • regulates ECM production and/or breakdown
  • maintains health of tissue
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10
Q

Anabolic cell molecules?

A
  • GF
  • anti-inflammatory cytokines
  • TIMPs
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11
Q

Catabolic cell molecules?

A
  • cytokines
  • MMPs
  • signaling mediators
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12
Q

What type of articular cartilage injuries occur in young and active population?

A
  • sporting and work related

- road traffic accidents

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13
Q

Non pharmalogical therapies for OA

A

FOR MILD SYMPTOMS

  • patient education
  • physical/occupational therapy
  • exercise and lifestyle
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14
Q

Therapeutic approached therapies for OA

A
  • NSAIDs
  • Rofecoxib
  • celecoxib (Pfizer)
  • Lumiracoxib
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15
Q

Disease modifying agents for OA

A
  • nutraceuticals (glucosamine)
  • DMOADs (cytokines, MMPs)
  • licofelone, COX
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16
Q

How to use autologous cell implantation?

A
  • health cartilage
  • isolate cells and expand
  • inject cells under periosteal graft
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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
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18
Q

Define mechanotransduction

A

activation of cell signalling

apply mechanical load to improve response of system/alter pathways and cell response

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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

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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
Q

Advanatges of compressive explant system

A
  • interactions of the matrix network are maintained in explants
26
Q

Disadvantages of compressive explant system

A
  • 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
Q

Examples of abnormal mechanical loading

A
  • altered joint loading
  • obesity
  • bone remodelling
  • trauma
28
Q

Examples of abnormal physiology

A
  • inflammation

- immune response

29
Q

Result of abnormal mechanical loading and abnormal physiology

A
  • matrix damage
  • abberant repair responses
  • mechanical failure
  • joint distraction, pain, disability
30
Q

In vivo events causing initiation of OA to early OA

A
  • mechanical injury
  • hereditary factors
  • ageing
31
Q

In vivo events causing early OA progression to late OA

A
  • inflammation
  • repetitive injury
  • subchondral bone changes
32
Q

What type of tissue organisation is cartilage?

A

Hetergenous

  • hyaline
  • chondrocytes
  • lacunae
  • matrix
  • collagen
  • proteoglycans
33
Q

What is the purpose of mechanical loading?

A
  • essential for normal metabolism
  • regulates ECM production/breakdown
  • maintains tissue health
  • biochemical and mechanical signals
34
Q

How can cartilage be repaired?

A

Total knee replacement

35
Q

What are some hypotheses on cell mechanotransduction?

A
  • 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
Q

How can we test the effect of dynamic compression?

A
  • 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
Q

What is the effect of dynamic compression in chondrocyte/agarose constructs?

A
  • 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
Q

How to test if response of chondrocyte/agarose constructs to dynamic compression is dependent on the cell sub-population?

A
  • 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
Q

Is the response of chondrocyte/agarose constructs to dynamic compressiondependent on the cell sub-population?

A
  • yes
  • superficial cells showed greater cell proliferation (mediated by NO and calcium)
  • deep cells showed greater proteoglycan synthesis (mediated by fluid flow)
40
Q

What are the different dynamic compression regimes and types?

A 
REGIMES
- continuous
- intermittent
TYPES
- unstrained
- strained
41
Q

What is the effect of continuous or intermittent compression on cell metabolism?

A
  • 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
Q

What are the principal receptors which percieve mechanical signals and activate signalling mediators?

A
  • integrins (a5B1)
43
Q

Which signalling mediators are involved in mechanotransduction process?

A
  • calcium ions

- NO

44
Q

How to test effect of TGFB on dynamic compression?

A
  • 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
Q

What is GRGDSP?

A

competitive ligand for the a5B1 integrin

46
Q

What is the effect of TGFB on dynamic compression?

A
  • 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
Q

How does IL-1B effect dynamic compression?

A
  • 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
Q

How does dynamic compression affect iNOS and COX-2 expression and production of NO and PGE2?

A

Our studies demonstrate an inhibition of iNOS and COX-2 expression and production of .NO and PGE2 release by dynamic compression.

Tissue Engineering & Regenerative Medicine (4 decks)

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