Nanog maintains human chondrocyte phenotype and function in vitro

Abstract

Previous work showed that Nanog, a homeobox family transcription factor, maintains embryonic stem cell pluripotency, suggesting that it has a role in stabilizing cell phenotype. Human chondrocytes lose their phenotype and dedifferentiate after relatively few passages in culture, changes that may limit their value in restoring damaged articular cartilage. We hypothesized that Nanog could stabilize the phenotype of cultured human chondrocytes in long-term monolayer cultures. To test this hypothesis, the human Nanog gene was stably transduced into human chondrocytes using a retroviral vector. Chondrocyte-specific gene expression (collagen type II, aggrecan, cartilage link protein, and Sox9) was measured by reverse transcription-polymerase chain reaction (RT-PCR) and real-time PCR in monolayer cultured chondrocytes transduced with Nanog and in control chondrocytes transduced with empty vector. In vitro cartilage matrix protein formation by Nanog-transduced and control cells was compared using Safranin-O and immunofluorescence stains. We found that after 25 passages, Nanog-transduced chondrocytes maintained significantly higher expression of collagen type II, aggrecan, and cartilage link protein genes than controls. Under chondrogenic conditions, Nanog-transduced cells produced significantly more cartilage-specific matrix than control cells. These findings support the hypothesis that Nanog maintains the human chondrocyte phenotype and function after long-term monolayer culture. Preservation of the chondrocyte phenotype may improve the ability of cultured chondrocytes to repair or restore articular cartilage.