Human fibroblast behavior on echinoderm-derived fibrillar collagen substrates

Soluble bovine collagen is one of the most used protein to prepare substrates and scaffolds for cell culture and biomedical applications mainly due to its high bio-compatibility. Nevertheless, it lacks the native fibrillar conformation typically found in the extracellular matrix. This feature reduces its bio-functionality and mechanical properties. Moreover, bovine collagen might increase the risk of prion disease transmission. Nowadays, new sources of collagen are welcome and the use of collagen extracted from aquatic organisms seems to be promising. In the present work, we extracted native fibrils from echinoderms, such as sea urchin, starfish and sea cucumber. Firstly, we optimized the protocol to specific body parts of these animals, then the extracted collagen was characterized in terms of ultrastructure and subsequently used to prepare fibrillar matrices that were deeply analyzed considering both ultrastructural and mechanical properties. All the obtained fibrils resembled the mammalian collagen mainly considering D-period and diameter. The matrix pore size was less than 2 µm, making impossible for a cell to pass through. In addition, the mechanical resistance (stiffness) was much higher (120-180 MPa) than that of soluble (mammalian) collagen substrates and comparable to values reported for the human skin. We also characterized the effect of these fibrillar collagen matrices on the organization of human skin-derived fibroblast cells (hSDF) as well as their substrate adhesion/interactions. Overall, hSDFs when seeded on fibrillar collagens appeared less numerous and with a more elongated shape than the controls (both bovine-derived collagen and plastic). The organization of F-actin appeared different, without stress fibers and with less numerous filopodial processes typical of the controls. These differences were probably due to the different substrate structural organization and mechanical properties, such as the stiffness, which need to be further investigated. All together, our data suggest that echinoderm fibrillar matrices could be an useful and promising tool for specific clinical/biomedical applications, such as artificial cell/tissue barriers with high mechanical resistance for Guided Tissue Regeneration (GTR), where a proper division between different anatomical compartments and no mixing up of different regenerating tissues and cytotypes are requested. Further in vivo studies will be carried out to confirm the bio-compatibility of these materials in alternative to bovine collagen matrices currently used for this kind of purpose.