Culture in Vitro of Human Osteoarthritis Chondrocytes

Objective: To study the separation,digestion and cultivation of human osteoarthritis chondrocytes,to compare biological characteristics with human normal chondrocytes and to evaluate their biological activity.Methods: The articular cartilage was digested one by one with the 0.05% and 0.2% collagenaseⅡ to separate chondrocytes in the culture medium including fetal bovine serum.The viability rate of the primary chondrocytes was detected.During the cultivation in vitro,the changes of the chondrocytes shape,growth and proliferation were observed;the changes of the collagen type Ⅱ and aggrecan were investigated and detected.Cell cycle,DNA ploidy and apoptosis of specimens in the presence or absence of IL-1β were evaluated by flow cytometry.Results: ① After the articular cartilage was completely dissolved by the enzymatic two-step digestion,the chondrocytes suspension were harvested.The viability rate of primary osteoarthritis chondrocytes was 82% on average,which was less than 95% of normal(P0.01).② The primary and passage osteoarthritis chondrocytes proliferated more slowly than the primary normal ones(P0.01).Their immunohistochemical staining of collagen type Ⅱ and heterochromia to toluidine blue O was both weaker than the normal.The fourth generation osteoarthritis chondrocytes had lost their special biological activity.③ The apoptotic rate of osteoarthritis chondrocytes was 6.9%,but that of normal was only 0.5%.After induced by IL-1β,the apoptotic rate of osteoarthritis chondrocytes increased to 27.4%,but that of normal increased to 12.7% only.Conclusion: The method of the enzymatic two-step digestion has advantages in the high cellular viability rate,low contamination and simple procedures.The osteoarthritis chondrocytes separated and cultured have biological characteristics of human osteoarthritis degenerated cartilage and can be regarded as the best experiment object for the research of osteoarthritis at the cell level.