Cell and material interactions in bone and cartilage tissue engineering

Due to their structural similarity to the native collagenous extracellular matrix (ECM), fibrous scaffolds are widely studied in cell-based cartilage and bone tissue engineering research. Recent studies report that cell behavior is affected by scaffold fiber diameter, although findings are inconsistent regarding whether fibers less than or greater than 1 micron are better substrates for chondrogenesis and osteogenesis. Differences in other experimental parameters between studies likely influence the outcomes, resulting in seemingly conflicting conclusions. Therefore, we explored the effects of scaffold fiber diameter on human mesenchymal stem cell (MSC) differentiation in conjunction with other culture parameters. Poly(e-caprolactone) (PCL) electrospun microfibers (3-4 m diameter) and nanofibers (300-400 nm diameter) were fabricated and utilized. First, we showed that microfibrous scaffolds are superior for inducing chondrogenesis at high initial seeding densities (2-4x10^6 cells/cm^3-scaffold), likely due to larger pore sizes within the scaffold promoting cell-cell contact as well as improved cellular infiltration. At lower seeding densities (1-5x10^5 cells/cm^3-scaffold) chondrogenesis is not strongly induced regardless of fiber diameter, suggesting high cell seeding densities are required to assess effects of biomaterials on chondrogenesis. Next, enhanced chondrogenesis on microfibers was found to be independent of tissue origin of MSCs, with much more robust chondrogenesis seen on microfibers than nanofibers for both bone marrow-derived MSCs (BMSCs) and adipose-derived MSCs (ADSCs), although the latter is generally less chondrogenic on either fiber diameter. Interestingly, under osteogenic conditions, BMSCs were not as sensitive to fiber diameter, but ADSCs demonstrated increased osteogenic potential on microfibers. Therefore, not only the lineage of differentiation, but also cell source must be taken into account when determining the optimal scaffold fiber diameter for cell-based skeletal tissue engineering. Lastly, we assessed the potential of enhancing MSC osteogenesis via sequential co-seeding with endothelial cells (ECs). Seeding MSCs onto scaffolds three days prior to ECs resulted in enhanced osteogenic differentiation in comparison to MSC monoculture, seeding ECs first, or simultaneously seeding both cell types. Taken together, these studies demonstrate that detailed examination of the interactions between different culture parameters, including scaffold architecture, seeding density, cell source, and differentiation lineage is essential for developing effective skeletal tissue engineering strategies