Mesenchymal stem cell activity across a graded scaffold-hydrogel composite biomaterial for tendon-to-bone enthesis repair

The severity of rotator cuff injury outcomes and a lack of tendon-to-bone enthesis regeneration strategies have inspired advances in biomaterials science to develop methods for interfacial tissue engineering. Here, we demonstrate a triphasic biomaterial comprising a non-mineralized, anisotropic collagen scaffold and a mineralized isotropic collagen scaffold linked via a continuous thiolated gelatin (Gel-SH) interface. This material provides a stratified environment in composition and porous architecture, and we report functional activity of human mesenchymal stem cells (hMSCs) across the scaffold. Notably, MSCs can be seeded onto the triphasic biomaterial and remain viable for up to 21 days. In addition, MSCs within the interfacial Gel-SH interfacial zone express markers associated with the rotator cuff fibrocartilaginous enthesis, including gene upregulation of <i>COL1A1, COL3A1, SOX9, BMP4, TGFβ1</i> and functional secretion of TGF-β1. Altogether, these findings suggest that this triphasic scaffold design could create a permissive environment for fibrochondrogenic activity in support of eventual enthesis interfacial tissue engineering applications.