Inspired by the aligned nanostructures and co-existence of vascular cells and stem cells in human cancellous bone, we quantitatively investigated the relative contributions of nanotopography and co-culture with human umbilical endothelial cells (HUVECs) to the osteogenesis of human mesenchymal stem cells (hMSCs). Although both nanotopography and co-culture independently enhanced the osteogenesis of hMSCs, osteogenesis was further enhanced by the two factors in combination, indicating the importance of synergistic cues in stem cell engineering. Interestingly, nanotopography provided a larger relative contribution to the osteogenesis of hMSCs than did co-culture with HUVECs. Furthermore, the osteogenesis of hMSCs was also affected by the density of parallel nanogrooves, exhibiting a maximum at a 1:3 spacing ratio, as defined as the ratio of ridge width to groove width. Analysis of (i) biochemical soluble factors, (ii) hMSC-substrate interaction and (iii) hMSC-HUVEC interaction suggests that (ii) and (iii) play a crucial role in mediating osteogenic phenotypes.
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