WEB Patterned collagen nanofibers induce morphological changes in fibroblasts
Nanotopographies control multiple cell functions such as adhesion, proliferation and morphology, as has been found in various studies with nanopatterns of synthetic polymers. However, in their native tissue environment cells are embedded into the nanofibrous extracellular matrix (ECM), which mainly consists of collagen. Yet, no protein scaffolds exist, which combine different topographies in the same substrate. Therefore, we established a new scaffold platform with nanofibrous and planar topographies in a single protein scaffold to study topography-related changes in cell proliferation and morphology.
Collagen scaffolds with nanofibrous and planar topographies were prepared by combining protein self-assembly with polymer patterning. The resulting binary collagen scaffolds were crosslinked with glutaraldehyde and the morphology of patterned collagen scaffolds was analyzed with scanning electron and atomic force microscopy. Subsequently, 3T3 mouse fibroblasts were cultivated on the binary collagen scaffolds for up to 72 hours, and cell proliferation and morphology were analyzed.
By combining polymer patterning with self-assembly of protein nanofibers we could fabricate collagen scaffolds with spatially controlled variations in the surface topography. Upon rehydration, nanofibrous collagen exhibited a surface roughness around 110 nm while planar collagen areas showed a roughness around 44 nm. When studying the growth of 3T3 fibroblasts for up to 72 hours we observed similar proliferation rates on nanofibrous and planar collagen. Interestingly, with confocal microscopy and SEM analysis we found pronounced differences in the fibroblast morphology in dependence of the underlying topography. On planar collagen fibroblasts exhibited a spread morphology with multiple short filopodia. On nanofibrous collagen areas fibroblasts were strongly elongated with a spindle-like shape and less filopodia. Overall, fibroblasts on planar collagen were larger than on collagen nanofibers.
Our results show that morphological changes in fibroblasts can be induced by spatially controlled variations in the surface topography of patterned collagen scaffolds. In the future, it will be highly interesting to transfer our patterning technique to the self-assembly of other fibrillar proteins like fibrinogen. In summary, our new scaffold design will allow us to track topography-dependent cell recognition processes on a single protein scaffold in real-time.