Enhancing Collagen Functionality with Bio-Stimulating Peptide Derivatization

Regenerative Medicine and Tissue Engineering are advanced fields of research dedicated to the development of novel approaches for the repair and regeneration of damaged or diseased tissues. One of the most promising strategies involves the use of natural biomaterials that mimic the properties of biological tissues, often modified to enhance their intrinsic characteristics. This study is part of a broader project focused on enhancing collagen scaffolds with specific signaling molecules to achieve targeted cell programming and direct desired actions within tissues and cells. In this pilot study, the goal is to improve collagen scaffold functionality through decoration with RGD, a bioactive peptide known for its ability to promote cell adhesion and osteogenicity. To achieve the binding between collagen and RGD, EDC was used as crosslinking agent following three different procedures: • P1: Collagen, EDC 1 mM, RGD 1 mM • P2: Collagen, MES 20 mM, EDC 1 M, RGD 1 mM • P3: Collagen, Acetic Acid 10 mM, MES 10 mM, EDC 0,5 M, RGD 0,5 mM. The bond formation was verified using several analytical techniques, including UV-VIS spectrophotometry, fluorimetry, and high-performance liquid chromatography (HPLC). Additionally, in vitro cellular assays with mesenchymal stem cells (MSCs) were conducted to evaluate the cytotoxicity, cell adhesion, and potential enhanced osteogenicity of RGD-decorated collagen. The results obtained from UV-VIS spectrophotometry, fluorimetry, and HPLC confirm that the collagen scaffold was successfully decorated with RGD using all three procedures, with the highest reaction yields obtained using P2 and P3. According to in vitro tests, scaffolds decorated following P2 and P3 were found to be cytotoxic, while the enhanced scaffold produced with P1 promotes cell adhesion, cell proliferation, and improves the osteogenicity of the collagen scaffold. Indeed, the P1 RGD-decorated scaffold exhibited ALP activity (an indicator of osteogenicity) that was 46.7% higher compared to undecorated collagen in standard DMEM media and 81.5% higher in osteogenic media.