Abstract
Objective
To extend the life span of human osteoarthritic (OA) articular chondrocytes by introduction of the catalytic component of human telomerase while preserving the chondrocyte‐specific phenotype.
Methods
Human articular chondrocytes were isolated from the femoral head and tibial plateau of patients undergoing knee joint replacement for OA. The chondrocytes were cultured as monolayers and infected with a retroviral telomerase expression construct followed by selection with G418 for 10–14 days. Telomeric‐repeat amplification protocol assays and telomere terminal restriction fragment length assays were performed on pools of transduced cells in order to measure telomerase activity and telomere length. Growth kinetics and population doubling capacity were assessed by passaging the cells in monolayer culture. Redifferentiation of the monolayer chondrocyte cultures was induced by transfer to suspension culture on poly‐(2‐hydroxyethyl‐methacrylate) (polyHEMA)–coated dishes. Induction of the chondrocyte‐specific phenotype was monitored by analysis of gene expression utilizing reverse transcription–polymerase chain reaction.
Results
OA chondrocytes isolated from 3 different donors (ages 41, 69, and 75 years) were transduced with a retroviral construct expressing telomerase. After selection, pooled populations of cells from all donors and a clonal cell line from 1 donor expressed telomerase activity and exhibited lengthening of telomeres. Chondrocytes expressing telomerase showed an increase of 5–9 population doublings over 234 days of culture in monolayer. The telomerase‐transduced cells recovered a chondrocyte‐specific gene expression pattern following culture on polyHEMA‐coated dishes.
Conclusion
The exogenous expression of telomerase may represent a way to expand human OA chondrocytes while allowing maintenance of the chondrocyte‐specific phenotype. These cells have the potential to be used for restoration of the articular cartilage defects occurring in this disease.