Identification of markers to characterize and sort human articular chondrocytes with enhanced in vitro chondrogenic capacity.
OBJECTIVE
To identify markers associated with the chondrogenic capacity of expanded human articular chondrocytes and to use these markers for sorting of more highly chondrogenic subpopulations.


METHODS
The chondrogenic capacity of chondrocyte populations derived from different donors (n = 21) or different clonal strains from the same cartilage biopsy specimen (n = 21) was defined based on the glycosaminoglycan (GAG) content of tissues generated using a pellet culture model. Selected cell populations were analyzed by microarray and flow cytometry. In some experiments, cells were sorted using antibodies against molecules found to be associated with differential chondrogenic capacity and again assessed in pellet cultures.


RESULTS
Significance Analysis of Microarrays indicated that chondrocytes with low chondrogenic capacity expressed higher levels of insulin-like growth factor 1 and of catabolic genes (e.g., matrix metalloproteinase 2, aggrecanase 2), while chondrocytes with high chondrogenic capacity expressed higher levels of genes involved in cell-cell or cell-matrix interactions (e.g., CD49c, CD49f). Flow cytometry analysis showed that CD44, CD151, and CD49c were expressed at significantly higher levels in chondrocytes with higher chondrogenic capacity. Flow cytometry analysis of clonal chondrocyte strains indicated that CD44 and CD151 could also identify more chondrogenic clones. Chondrocytes sorted for brighter CD49c or CD44 signal expression produced tissues with higher levels of GAG per DNA (up to 1.4-fold) and type II collagen messenger RNA (up to 3.4-fold) than did unsorted cells.


CONCLUSIONS
We identified markers that allow characterization of the capacity of monolayer-expanded chondrocytes to form in vitro cartilaginous tissue and enable enrichment for subpopulations with higher chondrogenic capacity. These markers might be used as a means to predict and possibly improve the outcome of cell-based cartilage repair techniques.
Grogan
Shawn Patrick
SP
University of Bern, Bern, Switzerland.
Barbero
Andrea
A
Diaz-Romero
Jose
J
Cleton-Jansen
Anne-Marie
AM
Soeder
Stephan
S
Whiteside
Robert
R
Hogendoorn
Pancras C W
PC
Farhadi
Jian
J
Aigner
Thomas
T
Martin
Ivan
I
Mainil-Varlet
Pierre
P
D051722:ADAM Proteins / Q000235:genetics / Q000378:metabolism; D000071116:ADAMTS5 Protein; D000328:Adult; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism; D000954:Antigens, Surface / Q000235:genetics / Q000378:metabolism*; D002358:Cartilage, Articular / Q000166:cytology / Q000378:metabolism*; D002478:Cells, Cultured; D019902:Chondrocytes / Q000145:classification* / Q000166:cytology / Q000378:metabolism*; D020219:Chondrogenesis / Q000235:genetics / Q000502:physiology*; D024043:Collagen Type II / Q000235:genetics / Q000378:metabolism; D005260:Female; D005434:Flow Cytometry / Q000379:methods; D006025:Glycosaminoglycans / Q000235:genetics / Q000378:metabolism; D006801:Humans; D018960:Hyaluronan Receptors / Q000235:genetics / Q000378:metabolism; D007334:Insulin-Like Growth Factor I / Q000235:genetics / Q000378:metabolism; D039422:Integrin alpha3 / Q000235:genetics / Q000378:metabolism; D008297:Male; D020778:Matrix Metalloproteinase 2 / Q000235:genetics / Q000378:metabolism; D008875:Middle Aged; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods; D060185:Tetraspanin 24
D015703:Antigens, CD; D000954:Antigens, Surface; C554715:CD151 protein, human; D024043:Collagen Type II; D006025:Glycosaminoglycans; D018960:Hyaluronan Receptors; D039422:Integrin alpha3; D060185:Tetraspanin 24; D007334:Insulin-Like Growth Factor I; D051722:ADAM Proteins; D000071116:ADAMTS5 Protein; C121532:ADAMTS5 protein, human; D020778:Matrix Metalloproteinase 2