关键词: 兔自体血清, 胎牛血清, 关节软骨细胞, 生物学特性, 反分化

Abstract:

BACKGROUND: At present, fetal bovine serum is mainly use for chondrocytes culture. However, because the security of the chondrocytes culture using heterologous serum in clinical application has been questioned in recent years, the autologous serum for chondrocytes culture has been paid more and more attention.
OBJECTIVE: To compare the difference in the biological properties between rabbit articular chondrocytes cultured with 10% autologous serum and 10% fetal bovine serum in vitro.
METHODS: After the culture medium of rat autologous serum was prepared, rabbit articular chondrocytes were isolated from rabbits and seeded into the medium with 10% autologous serum and 10% fetal bovine serum, respectively for monolayer subcultivation. Then, passage1, 3 and 5 cells were chosen and cell morphological changes were observed by light microscope, meanwhile, growth curves were drawn to evaluate cell poliferation rate. Besides, biological properties of chondrocytes were observed by toluidine blue staining, typeⅠ and typeⅡ collagen immunohistochemical staining. In addition, the expression changes of typeⅠ, typeⅡcollagen, CD26 and CD44 were analyzed by flow cytometry.
RESULTS AND CONCLUSION: (1)There was no significant difference in cell morphology between chondrocytes cultured with autologous serum and fetal bovine serum. (2)The cell growth rate of chondrocytes cultured with autologous serum was m ore rapid than that of chondrocytes cultured with fetal bovine serum. (3)Toluidine blue staining showed that both chondrocytes cultured with autologous serum and fetal bovine serum showed a passage-dependent reduction of staining. In passage 5 cells, the two groups almost had no different dying. For passage 1 and 3 cells, chondrocytes cultured with autologous serum showed deeper staining than fetal bovine serum. (4)Immunohistochemical staining and flow cytometry results showed that expression of type Ⅰ collagen was increased with passage, while type Ⅱ collagen expression was decreased with passage. Moreover, typeⅠcollagen expression of chondrocytes cultured with autologous serum was significantly lower than that chondrocytes cultured with fetal bovine serum at passage 3 (P < 0.05). (5)CD26 expression was increased at first and then decreased, while the expression of CD44 did not change with passage. These results suggest that compared with 10% fetal bovine serum, the growth rate of chondrocytes cultured with 10% autologous serum is more rapid, and has a better preservation of phenotype based on the results of different observations.