Fabrication of injectable, cellular, anisotropic collagen tissue equivalents with modular fibrillar densities

Abstract Technological improvements in collagen gel fabrication are highly desirable as they may enable significant advances in the formation of tissue-equivalent biomaterials for regenerative medicine, three-dimensional (3D) in vitro tissue models, and injectable scaffolds for cell and drug delivery applications. Thus, strategies to modulate collagen gel fibrillar density and organization in the mesostructure have been pursued to fabricate collagenous matrices with extracellular matrix-like features. Herein, we introduce a robust and simple method, namely gel aspiration-ejection (GAE), to engineer 3D, anisotropic, cell seeded, injectable dense collagen (I-DC) gels with controllable fibrillar densities, without the use of crosslinking. GAE allows for the hybridization of collagen gels with bioactive agents for increased functionality and supports highly aligned homogenous cell seeding, thus providing I-DC gels with distinct properties when compared to isotropic DC gels of random fibrillar orientation. The hybridization of I-DC with anionic fibroin derived polypeptides resulted in the nucleation of carbonated hydroxyapatite within the aligned nanofibrillar network upon exposure to simulated body fluid, yielding a 3D, anisotropic, mineralized collagen matrix. In addition, I-DC gels accelerated the osteoblastic differentiation of seeded murine mesenchymal stem cells (m-MSCs) when exposed to osteogenic supplements, which resulted in the cell-mediated, bulk mineralization of the osteoid-like gels. In addition, and upon exposure to neuronal transdifferentiation medium, I-DC gels supported and accelerated the differentiation of m-MSCs toward neuronal cells. In conclusion, collagen GAE presents interesting opportunities in a number of fields spanning tissue engineering and regenerative medicine to drug and cell delivery.