Multi-Level Factorial Analysis of Ca2+/Pi Supplementation as Bio-Instructive Media for In Vitro Biomimetic Engineering of Three-Dimensional Osteogenic Hybrids

We report on the in vitro use of Ca2+/P-i supplementation as a bio-instructive medium to drive human periosteum-derived cells (hPDCs) toward osteogenic differentiation on three-dimensional (3D) porous Ti6A14V scaffolds. Through a multilevel factorial analysis, we have systematically investigated the biological effect and interactions of Ca2+ or P-i supplementation in three selected media preparations (i.e., basic growth medium, osteogenic medium [OM, and osteogenic medium without beta-glycerophosphate [OM-]) and have identified specific conditions which induce proliferation and significant osteogenic differentiation of two-dimensional (2D) hPDC cultures. These findings were translated from 2D to 3D cultures conditions to instruct hPDCs to populate porous Ti6A14V scaffolds and to differentiate into the osteoblast lineage with collagenous matrix production and subsequent matrix mineralization on the 3D structures. These osteogenic hybrids may potentially serve as a clinically relevant customizable bone reparative unit, providing a biomimetic template to more effectively mediate in vivo bone regeneration.