Evaluation of cartilage specific matrix synthesis of human articular chondrocytes after extended propagation on microcarriers by image analysis.
BACKGROUND
Cell-based technologies for the repair of cartilage defects usually rely on the expansion of low numbers of chondrocytes isolated from biopsies of healthy cartilage. Proliferating chondrocytes are known to undergo dedifferentiation characterized by downregulation of collagen type II and proteoglycan production, and by upregulation of collagen type I synthesis. Re-expression of cartilage specific matrix components by expanded chondrocytes is therefore critical for successful cartilage repair.


METHODS
Human articular chondrocytes were expanded on microcarriers Cytodex 3. The growth area was increased by adding empty microcarriers. Added microcarriers were colonized by bead-to-bead transfer of the cells. The chondrocytes were harvested from the microcarriers and characterized by their ability to synthesize collagen type II when cultivated in alginate beads using chondrogenic growth factors. A semi-automatic image analysis technique was developed to determine the fractions of collagen type II and type I positive cells.


RESULTS
The expansion of human articular chondrocytes on microcarriers yielded high cell numbers and propagation rates compared to chondrocytes expanded in flask culture for one passage. The proportion of collagen type II positive cells compared to collagen type I synthesizing cells was increased compared to chondrocytes expanded using conventional methods. The matrix synthesis upon treatment with chondrogenic factors IGF-I and BMP-7 was enhanced whereas TGF-ss had an inhibitory effect on microcarrier expanded chondrocytes.


CONCLUSIONS
Expanding human articular chondrocytes on microcarriers omitting subcultivation steps leads to superior ratios of collagen type II to type I forming cells compared to the expansion in conventional monolayer culture.
Goepfert
Christiane
C
Hamburg University of Technology, Institute of Bioprocess and Biosystems Engineering, Hamburg - Germany.
Lutz
Vivien
V
Lünse
Svenja
S
Kittel
Sabrina
S
Wiegandt
Katharina
K
Kammal
Michael
M
Püschel
Klaus
K
Pörtner
Ralf
R
D055419:Bone Morphogenetic Protein 7 / Q000494:pharmacology; D002358:Cartilage, Articular / Q000166:cytology*; D002454:Cell Differentiation / Q000502:physiology; D019902:Chondrocytes / Q000378:metabolism*; D003094:Collagen / Q000187:drug effects / Q000378:metabolism*; D024043:Collagen Type II / Q000378:metabolism; D006801:Humans; D007091:Image Processing, Computer-Assisted; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology; D023822:Tissue Engineering / Q000379:methods*; D016212:Transforming Growth Factor beta / Q000494:pharmacology
D055419:Bone Morphogenetic Protein 7; D024043:Collagen Type II; D016212:Transforming Growth Factor beta; D007334:Insulin-Like Growth Factor I; D003094:Collagen