Biomimetic bone regeneration using angle-ply collagen membrane-supported cell sheets subjected to mechanical conditioning.

Bone injuries are common and new strategies are desired for achieving ideal bone regeneration for bone defect repair. Scaffolds with bone-mimicking characteristics may provide an appropriate microenvironment to promote bone regeneration. Meanwhile, mechanical stimulation effectively regulates a wide range of cellular behaviors such as cell proliferation and differentiation. In this study, biomimetic multi-layer cell-collagen constructs with angle-ply structural feature were prepared by assembling micropatterned collagen membranes on which aligned MC3T3-E1 cells were cultured. The anisotropic microgrooved collagen membranes effectively guided the alignment of cells and promoted the osteogenic differentiation of them. To further promote cell differentiation and extracellular matrix production, the multi-layer cell-collagen constructs were cultured under mechanical conditioning through cyclic stretching. It was found that the constructs with both cell alignment and mechanical conditioning resulted in better osteogenic potential than those with cell alignment or mechanical conditioning alone. Upon implantation into the critical-sized calvarial defects of mice, the constructs with both cell alignment and mechanical conditioning achieved best new bone formation efficacy. Together, findings from this study reveal that synergized use of microstructural and mechanical cues may provide an effective new approach toward bone regeneration. STATEMENT OF SIGNIFICANCE: Biomimicking is an effective strategy to promote bone regeneration for repairing bone defects. Although numerous studies which micro-structurally mimicked native bone using various scaffolds, far less studies have paid attention to the mechanical environment of bone. In this study, angle-ply collagen membrane-supported cell sheets were prepared and pre-conditioned using mechanical loading prior to implantation at bone defects. The constructs with cell alignment and subjected to mechanical conditioning resulted in better osteogenic differentiation of cells in vitro and new bone formation in vivo than those with cell alignment or mechanical conditioning alone. Therefore, recapitulation of both microstructural and mechanical features of native bone may result in a synergistic effect and provides an effective approach toward bone regeneration.