Copolymer cell/scaffold constructs for bone tissue engineering: co-culture of low ratios of human endothelial and osteoblast-like cells in a dynamic culture system.

The aim of this study was to compare the effect of different ratios of human umbilical vein endothelial cells (HUVECs) on osteogenic activity of human osteoblast-like cells (HOB) and capillary-like structure (CLS), seeded into copolymer scaffolds in a dynamic culture system. HOB and HUVEC were co-cultured into poly(L-lactide)-co-(1,5-dioxepan-2-one) [poly(LLA-co-DXO)] scaffolds at ratios of 5:1 (5:1 group) and 2:1 (2:1 group). Samples were collected after 5, 15, and 25 days. Cross-sections were processed and the CLS from HUVEC was disclosed in both groups. Cell viability was determined by dsDNA assay. Cells seeded at the ratio of 5:1 had good viability. Total RNA was isolated and the reverse transcription reaction was performed. The influences on the expression of several osteogenic genes were various with regarding to different ratios of HUVEC demonstrated by the PCR array. The RT-PCR results was in consistent with the PCR array results that several osteogenesis related genes had higher expression in the 5:1 group than in the 2:1 group, especially at day 25, such as alkaline phosphatase, insulin-like growth factor 1 (IGF1), and so forth. ELISA showed that the production of IGF1 after 25 days of incubation were higher in cells co-cultured at the 5:1 ratio than at the 2:1 ratio. The results show that under dynamic culture conditions, co-culture of HOB with a low ratio of HUVEC in copolymer scaffolds results in CLS formation and significantly influenced the expression of osteogenic markers.