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Tissue Engineeringof
CartilageEvan WitmerGregory Lynn
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Cartilage Basics
Ailments, Traditional Treatments, &Tissue Engineering
Six Current Publications Objective & Justification Methods & Materials Results
Summary
Acknowledgements
Overview
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What is Cartilage?
Gel-like connective tissue made
up of chondrocytes, collagen,proteoglycans, and other ECM
proteins
70-80%water
Avascular
no nutrients, no regeneration
3 types:
Hyaline (low-friction)
Elastic (epiglottis)
Fibrocartilage (shock absorbing)
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Ailments & Traditional Treatme
Causes: Trauma, chronic wea
developmental disorders, im
Little-to-no natural regen Autografts (self) / Allogr
transplantation of do Debridement (smoothing Marrow stimulation
Problems: site morbidity, lim
availability, immune responsPrimary Source: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3146065/
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Advent ofTissue Engineerin
Creates an embryonic-like environment that stimulates grow Custom-made scaffolds for strength, structure, and function Resulting structure is true self-regenerated tissue Single surgery required
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Objective: To better assess the applicability of the electrospinnintechnology for scaffold fabrication, through electrospinning six co(-hydroxy esters) and testing their characteristics
Electrospinning high surface area to volume ratio similar structural morphology to the fibrillar ECM
Poly(-hydroxy esters) biodegradable & FDA approved
IF: 5.68 - (200
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Methods & Materials
6 Poly(-hydroxy esters)spun into fibrous scaffolds
A.) PGAB.) PLGA5050
C.) PDLLA
D.)PLGA8515
E.) PLLA
F.) PCL
Analysis Coat in gold & SEM Apply load
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Results
Cell-Matrix Interaction (After 7 days) Remained on surface of PGA, PDLLA, PLGA5050, and PL
Chondrocytes integrated into PLLA & PCL Cellular proliferation
Chondrocytes Mesenchymal Stem Ce
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(201Objective: To describe the developmental physicochemical pro
silk fibroin scaffolds derived from high-concentrationaqueousolutions.
Silk fibroin in vivo degrades easily tailored & processed
More than 10% aqueous silk fibroin solution
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Methods & Materials
Silk fibroin scaffolds with
different initialconcentrations(in wt.%)
A.) 8%
C.) 10%
E.) 12%
G.) 16%,
Combined methodology Salt-leaching & freeze-
drying Create different
structures
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Results
All above 90% interconnectivityand79% porosity
~15% of pores larger than 300m Water up-take (30 days immersion)
Original weights No significant differencein
morphology Mechanical Properties
Silk-16& Silk-12values suitablefor meniscus and cartilageengineering
Compressive
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European Cel(2008)
Objective: To evaluate in vivo cartilaginous tissue formation by cseeded fibrin/PLGA hybrid scaffolds.
Joint replacement PGLA & fibrin scaffolds
in vitro promoted cartilage constructs
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Methods & Materials
fibrin/PLGA hybrid (A) PLGA scaffolds soaked in
chondrocyte-fibrin solution
Control: Chondrocytes seededintojust PLGA (B)
Implanted into dorsum of nudemice 4 weeks
A
B
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Results
Fibrin/PLGA hybrid superior Higher cell viability and
proliferation Superior cell distribution, cell-
matrix organization, overallECM production
Fibrin/PLGA
PLGA
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Biomac(2005
Objective: To combine the benefits of a photocrosslinkable netwthe desirable material HA for cartilage tissue engineering.
Hyaluronic Acid (HA) forms aggrecans (proteoglycan CSPCP) &
wound healing Hydrogels photoencapsulate chondrocytes
Easier to fill tight spaces and irregularly shaped defects
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Materials & Methods
Materials:
HA and methacrylic anhydrideform MeHA Polyethylene glycol
dimethacrylate (PEGDM) Swine chondrocytes Nude mice
Methods: Fabrication of MeHA & PEGDM Encapsulation by polymerizing with UV light Culture in nude mice 4, 6, and 8 week dissection
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Results
Mechanical properties linear slope at low strains
(
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Results Cell viability
Decrease in viability with increase inmonomer concentration
Effect of molecular weight wasnegligible
1 day (black) 1 w
immediate (black), 4 weeks (gray), 8 weeks (white)
Neocartilage formation After 4 weeks, all co
became opaque. StaGAG formation
As effective as the Pbased hydrogel
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(200IF: 8.
Objective: Study aims to form non-toxic composite and study meproperties and degradation of chitosan-HAin vitro.
New Material: water-soluble chitosan and oxidized hyaluroni
without toxic cross-linking Crosslinking action with amino functions
No photopolymerization to prevent prolonged irradiation, nophotosensitizer (toxic)
h d i l
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Methods & Materials
Materials: Chitosan
HA sodium Succinic anhydride Bovine chondrocytes
Methods Succinyl-CS and Aldehyde-HA
synthesized and lyophilized
Hydrogels formed by mixing ratios of1:9, 3:7, 7:3, 9:1
Schiffs base crosslinking reaction Encapsulation:
Mix cells with S-CS, Add A-HA to form gel
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Results
Cell adhesion 5:5 & 7:3 signif
greater than 3
Degradation rate strong correlation to ratio of S-
CS to A-HA more S-CS = slower weight loss
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Results
Cell proliferation and viability successful encapsulation
in a normal sphericalmorphology
promoted cell survival
Successful crospossible with mreagents
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(2004)IF: 8.31
Objective: To study the response of bovine chondrocytes on natichemically modified BC (Phosphorylation & Sulfation)
Bacterial cellulose (BC) has unusual material properties & deg
high water retention, fiber network, high wet tensile stre cost-effective mass producible BC mold
Lack of literature describing native mechanical properties forconstructs
M th d & M t i l
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Methods & Materials Materials
G. xylinus H
3PO
4
NH2SO3H Bovine chondrocytes
Methods G. xylinus grown & BCpurified Phosphorylation
BC-P1 (30 min), BC-P2 (2 hr)
Sulfation BC-S
Cultured human arterial cartilage for 8days collagen type II, plant-derived
cellulose, calcium alginate, and
tissue culture plastic
l
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Results
Modifications changedsurface morphology to
adhere more strongly
Strong immune responseto native BC
Modified BC showshigher growth at similarimmune response toalginate & tissue cultureplastic
R lt
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Results
Presence of cellingrowthpromisingfor scaffoldmaterialengineering
BC does notprematurelydifferentiate cellsto form fibroblasts
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Summary
Scaffold Materials:
Synthetics PGA, PLLA, PDLLA,
PLGA5050, PLGA8515, PCL Natural
Silk, Bacterial Cellulose, HA,Chitosan
Huge variety of materials
Techniques:
Electrospinning,Lyophilization,Injectable HydrogPhotopolymeriza
Cross-linkinggenerally determdegradation rate
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Future of Cartilage TE
Hydrogels mimicking ECM-like matrices Self-assembling nanoscaffolds Thermosensitive injectable materials Biomimetic novel materials (coral, silk, etc.)
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Acknowledgements
Dr. Abidian
Evan Witmer
Greg Lynn