Cartilage tissue engineering by electrospun PCL scaffolds seeded with rabbit chondrocytes under flow perfusion culture in vitro / 中华骨科杂志

ABSTRACT

Objective To investigate the chondrocyte proliferation and extracellular matrix biosynthesis of electrospun PCL scaffolds seeded with rabbit chondrocytes under flow perfusion culture in vitro.Methods Nonwoven PCL microfiber mats were fabricated,and contra-aperture cylindrical glass equipment as a perfusion bioreactor was designed and manufactured on our own.The experiment included peffusion culture group and static culture group.Primary chondrocytes were isolated from the knee joints of two-month-old New Zealand white rabbits and seeded into scaffolds.The scaffold-cell complexes were harvested at 3,7,and 14 days of culture for scanning electron micrograph (SEM) analysis,biochemical assay,real-time PCR and histology analysis.Results Electrospun PCL scaffolds were composed of microfibers with a diameter of 1.67±0.76 μm and pores with a diameter of 17.65±7.11 μm.SEM showed a better cell proliferation with typical morphology of chondrocytes under perfusion culture.At 7 days of culture,DNA content in perfusion culture group was higher than in static culture group.At 3,7 and 14 days of culture,compared with the static culture group,glycosaminoglycan (GAG) content and GAG/DNA ratio in perfusion culture group were higher,and the differences were statistically significant.At 14 days of culture,real-time PCR showed aggrecan and collagen type Ⅱ gene expression and collagen type Ⅱ to collagen type Ⅰ ratio were higher in perfusion culture group than in static culture group; HE and safranin O staining showed a significant cell proliferation,infiltration,as well as extracellular matrix biosynthesis in perfusion culture group.Conclusion Under flow perfusion culture,the electrospun PCL scaffolds seeded with rabbit chondrocytes can enhance chondrocyte proliferation and extracellular matrix biosynthesis,which is a promising method for cartilage tissue engineering.