Macro-Scaled M13 Bacteriophage-Based Hierarchical Fiber for Piezoelectricity-Induced Cell Regeneration

Regenerative medicine is a major domain of medical technology that aims to restore and develop tissue or organs’ own functions by replacing or regenerating damaged human cells, tissues, and organs. Of these targets, tissue engineering is a promising method used to identify damage from aging, injury, and degenerative diseases by reestablishing structural and functional properties of tissues, or organs. Previously, scaffolds produced with biocompatible materials have been developed that stimulate tissue regeneration at the cellular or molecular level. However, most of the research regarding scaffolds with biomaterials focused on mechanical support and biofunctionality, which is related to the enhancement of biological interactions with bodily components. These interactions provoke stimulatory actions, although, the effect is insufficient to regenerate the tissue of interest and to make a clinical trial. In this regard, we proposed a system that produces microcurrents, relatively weak currents that mimic the spontaneous generation of currents in the human body, for tissue-specific generation and promotion of cell development/regeneration. These microcurrents induce a therapeutic effect that treats damaged bodily tissues retaining piezoelectric coefficients. The system is composed of an M13 bacteriophage-based material that was hierarchically self-assembled to create a fibrous structure using a bottom-up strategy. A magnetic field was applied to M13 phage solution for unidirectional alignment of the material, followed by fixation. The optical anisotropic property of the aligned macrostructure was qualitatively analyzed using polarized optical microscopy, and quantitatively analyzed using birefringence measurements. When sample was placed between the polarizer and analyzer, there were difference of image appeared between the highly aligned structure and randomly aligned structure. The highly aligned structure presented a higher birefringence value (Δn=|<span style="font-size:10.8333px">n_e-n_o|</span>) than randomly aligned structure that was not applied magnetic field. Furthermore, Unidirectional alignment of the structure was qualitatively, and quantitatively confirmed through Second Harmonic Generation (SHG) microscopy and its SHG intensity due to the non-linear polarization induced by interactions between photons of incident electromagnetic waves and electrons of molecules constituting non-centrosymmetrical material. The piezoelectric coefficient of the structure can be increased by unidirectionally aligning the bio-piezoelectric M13 phages, which can be adjusted depending on the concentration of the phage and intensity of the applied magnetic field to mimic the piezoelectric coefficient of the target tissue. Specifically, we investigated the piezoelectric response of osteoblasts to bone piezoelectricity-mimetic material. When mechanical force was applied, the phage-based macrostructural material generated a piezoelectric response specific to the bone and promoted proliferation and differentiation of the osteoblasts to osteocytes in vitro. To the best of our knowledge, this is the first case in which biomolecules of a single type are assembled to fabricate a piezoelectric macrostructure that produces a piezoelectric coefficient comparable to signals from bodily tissues. We plan to further develop the system to characterize effects of 3-dimensional scaffolds that made up of M13 phage-based hierarchical materials into various cells or tissues in vitro as well as in vivo.