Retroviral transduction with SOX9 enhances re-expression of the chondrocyte phenotype in passaged osteoarthritic human articular chondrocytes.
OBJECTIVE
Articular chondrocytes proliferate in monolayer culture, but the expression of the transcription factor SOX9 falls and the ability of the cells to reform cartilage tissue declines. We have investigated whether retroviral SOX9 expression in extensively passaged human articular chondrocytes from osteoarthritic (OA) joints enables the cells to regain a cartilage matrix forming phenotype in pellet culture.


METHODS
Chondrocytes from normal and OA joints were retrovirally transduced with SOX9 and grown to passages 7-10 before being cultured as pellets of 500,000 cells for 14 days. Pellets were analysed by real time polymerase chain reaction, histology, immunohistochemistry and 1,9-dimethylmethylene blue assay.


RESULTS
Chondrocytes from OA joints displayed higher expression of COL2A1 gene when transduced with SOX9 and cultured as pellets with 10% serum, but glycosaminoglycan (GAG) synthesis was low. Addition of transforming growth factor beta-3 and insulin like growth factor-1 increased collagen II expression and GAG synthesis in these SOX9 transduced cell pellets. The cells adopted a rounded morphology and there was increased deposition of collagen II protein compared to control green fluorescent protein transduced cell pellets. Similar results were seen with transduced chondrocytes from OA or healthy cartilage. SOX9 transduced human dermal fibroblasts did not show any chondrogenic response.


CONCLUSIONS
Transduction with SOX9 primed the passaged articular chondrocytes to regain a chondrocytic phenotype in pellet culture and to form a cartilaginous matrix, which was enhanced by growth factors. Following transduction, chondrocytes from OA joints showed a similar capacity for chondrogenic recovery as those from healthy joints, which suggested that OA does not permanently compromise the chondrocyte phenotype.
Tew
Simon R
SR
UK Centre for Tissue Engineering at The Wellcome Trust Centre for Cell-Matrix Research, Faculty of Life Sciences, University of Manchester, Manchester, UK.
Li
Ying
Y
Pothacharoen
Peraphan
P
Tweats
Lisa M
LM
Hawkins
Robert E
RE
Hardingham
Timothy E
TE
D002358:Cartilage, Articular / Q000378:metabolism* / Q000473:pathology; D018929:Cell Culture Techniques; D019902:Chondrocytes / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D024043:Collagen Type II / Q000378:metabolism; D005109:Extracellular Matrix / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D005822:Genetic Vectors; D006025:Glycosaminoglycans / Q000096:biosynthesis; D006609:High Mobility Group Proteins / Q000235:genetics / Q000502:physiology*; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology; D020370:Osteoarthritis, Knee / Q000378:metabolism / Q000473:pathology*; D010641:Phenotype; D012190:Retroviridae / Q000235:genetics; D055758:SOX9 Transcription Factor; D012867:Skin / Q000166:cytology / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology*; D014161:Transduction, Genetic; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D053782:Transforming Growth Factor beta3
D024043:Collagen Type II; D006025:Glycosaminoglycans; D006609:High Mobility Group Proteins; D055758:SOX9 Transcription Factor; C527932:SOX9 protein, human; D014157:Transcription Factors; D016212:Transforming Growth Factor beta; D053782:Transforming Growth Factor beta3; D007334:Insulin-Like Growth Factor I