Rabbit articular chondrocyte dedifferentiation during in vitro expansion

doi:10.3969/j.issn.2095-4344.2013.20.003

Abstract

Abstract:

BACKGROUND: Rabbits have been extensively utilized in cartilage tissue engineering as experimental models. However, research efforts remain limited concerning the dedifferentiation of rabbit articular chondrocytes.
OBJECTIVE: To explore the dedifferentiation of rabbit articular chondrocytes during in vitro expansion.
METHODS: Chondrocytes were isolated from articular cartilage of New Zealand white rabbits and subcultured in vitro until passage 7. Cells were analyzed regarding cell growth, morphology, deposition of cartilaginous extracellular matrix and gene expression using cells counting method, microscopic examination, F-actin staining, safranine-O staining, quantitative determination of glycosaminoglycans and semiquantitative reverse transcription-polymerase chain reaction, respectively
RESULTS AND CONCLUSION: Under light microscope, rabbit articular chondrocytes changed from small round or polygonal shape into fibroblast-like morphology during the in vitro subculture, which was further confirmed with F-actin staining using phalloidin. Results of cell counting showed that, cell proliferation significantly declined with passage generations, especially after passage 3, articular chondrocytes were not proliferating. Safranine-O staining and glycosaminoglycans quantification demonstrated that, the capacity of producing cartilaginous extracellular matrix decreased dramatically as early as passage 2. Semiquantitative polymerase chain reaction analysis showed that, the gene expression of collagen Ⅱ, aggrecan, cartilage oligomeric matrix protein and SOX9 was downregulated especially for cells after passage 3, while the expression of collagen I and versican was upregulated. In addition, the CD90 expression increased and CD14 expression remained unaltered. Rabbit articular chondrocytes dedifferentiate quickly upon in vitro expansion with distinct variation of gene expression profile, and cells within passage 3 are appropriate for applications in cartilage repair.

Key words: tissue construction, cartilage tissue construction, articular cartilage damage, cartilage tissue engineering, rabbit, chondrocytes, in vitro expansion, dedifferentiation, cell morphology, cell growth, histological staining, gene expression, ministerial grants-supported paper

CLC Number:

R318

Xu Lei, Ye Zhao-yang, Zhou Yan, Tan Wen-song. Rabbit articular chondrocyte dedifferentiation during in vitro expansion [J]. Chinese Journal of Tissue Engineering Research, 2013, 17(20): 3626-3634.

Figures/Tables 8

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