Short Poster Lecture
Biomaterials featuring patterns in mechanical properties and biomolecule presentation hold great promise for tissue engineering applications, as they could enable spatial control over cell behavior and tissue formation. We developed an alginate-based material system that permits the creation of these patterns on the µm-scale by combining two orthogonal covalent crosslinking chemistries: Diels-Alder and thiol-ene. As the Diels-Alder reaction occurs spontaneously and the thiol-ene reaction requires initiation by UV, the precise location and timing of this second step is controlled via photomasks to generate patterns.
Initial experiments were performed on single-crosslinked, unpatterned materials. Diels-Alder crosslinked hydrogels were fabricated by combining alginate modified with norbornene or tetrazine functional groups. The degree of crosslinking and resulting mechanical properties were controlled by the amount of norbornene and the stoichiometric ratio of norbornene to tetrazine. Thiol-ene crosslinked hydrogels were prepared by exposing norbornene-coupled alginate and a dithiol crosslinker to UV, and mechanical properties were tuned by changing the amount of crosslinker. Both material sets were characterized for rheological and mechanical properties.
Materials were rendered cell-compatible via conjugation of thiol-coupled RGD sequences linked to remaining available norbornene groups. This modification was confirmed by quantifying viability and proliferation of mouse pre-osteoblasts in 2D and 3D over one week.
While preparing dual-crosslinked, single-phase materials, it was noted that the timing of UV exposure (and therefore thiol-ene crosslinking) enables control over stiffness: instant UV exposure led to soft gels, while later UV exposure led to increased gel stiffness.
Patterns in mechanical properties were confirmed by nanoindentation surface mapping. Patterns in RGD molecule and BMP-2 peptide mimic presentation were visualized by subsequent mesenchymal stromal cell (MSC) attachment onto modified regions and quantifying cell morphology or differentiation into the osteogenic lineage, respectively.
This biomaterial-based strategy allowing biophysical and biochemical patterning could direct MSC behavior and support guided tissue regeneration in clinically-challenging situations.