Bone tissue, by definition, is an organic–inorganic nanocomposite, where metabolically active cells are embedded within a matrix that is heavily calcified on the nanoscale. Currently, there are no strategies that replicate these definitive characteristics of bone tissue. Here we describe a biomimetic approach where a supersaturated calcium and phosphate medium is used in combination with a non-collagenous protein analog to direct the deposition of nanoscale apatite, both in the intra- and extrafibrillar spaces of collagen embedded with osteoprogenitor, vascular, and neural cells. This process enables engineering of bone models replicating the key hallmarks of the bone cellular and extracellular microenvironment, including its protein-guided biomineralization, nanostructure, vasculature, innervation, inherent osteoinductive properties (without exogenous supplements), and cell-homing effects on bone-targeting diseases, such as prostate cancer. Ultimately, this approach enables fabrication of bone-like tissue models with high levels of biomimicry that may have broad implications for disease modeling, drug discovery, and regenerative engineering.

Microbially induced calcite precipitation (MICP) was used to treat several sandboxes filled with naturally occurring beach sand collected from Atlantic Beach, Florida. A surface-spray/percolation technique was used to treat these sandboxes where a relatively high-concentration bacteria solution and high-concentration calcium chloride/urea solutions were applied directly to the boxes’ surfaces. Several different treatment combinations were tested whereby bacterial optical density, bacteria/urea/calcium chloride volume relative to pore volume, and bacteria/urea/calcium chloride ratio were manipulated. Treated sandboxes were tested for erodibility using a pocket erodometer. In addition, sandboxes were dissected after erosion testing to examine crust depth. Results showed that higher optical densities, higher bacteria quantities relative to void volume, and higher bacteria quantities relative to urea led to lower erodibility and greater crust depth. When MICP constituent quantities were maximized to give the best erosion/crust-depth results, erodibility improvements began to approach what may be considered adequate erosive resistance. Further investigation is required to better classify this behavior more quantitatively.

The hierarchical structure of bone and intrinsic material properties of its two primary constituents, carbonated apatite and fibrillar collagen, when being synergistically organized into an interpenetrating hard-soft composite, contribute to its excellent mechanical properties. Lamellar bone is the predominant structural motif in mammalian hard tissues; therefore, we believe the fabrication of a collagen/apatite composite with a hierarchical structure that emulates bone, consisting of a dense lamellar microstructure and a mineralized collagen fibril nanostructure, is an important first step toward the goal of regenerative bone tissue engineering. In this work, we exploit the liquid crystalline properties of collagen to fabricate dense matrices that assemble with cholesteric organization. The matrices were crosslinked via carbodiimide chemistry to improve mechanical properties, and are subsequently mineralized via the polymer-induced liquid-precursor (PILP) process to promote intrafibrillar mineralization. Neither the crosslinking procedure nor the mineralization affected the cholesteric collagen microstructures; notably, there was a positive trend toward higher stiffness with increasing crosslink density when measured by cantilever-based atomic force microscopy (AFM) nanoindentation. In the dry state, the average moduli of moderately (X51; 4.8 ± 4.3 GPa) and highly (X76; 7.8 ± 6.7 GPa) crosslinked PILP-mineralized liquid crystalline collagen (LCC) scaffolds were higher than the average modulus of bovine bone (5.5 ± 5.6 GPa).