SpinoMimetic CryoSponges as advanced biomaterial archetypes for enhanced neuronal anchoring and support Achievement of maximal mechanotransduction via minimal functionalization and crosslinking

摘要

Introduction: Mechanical properties of a biomaterial scaffold play a definitive role in neuronal support, differentiation and even regeneration. To achieve neuromimetism, mechanotransduction through focused selection of biomaterials (including blends) require fine-tuning of their chemical morphology In this study, novel "SpinoMimetic CryoSponge Architectures" were developed employing various polyethylene glycol (PEG) derivatives (2kDa) with chitosan network for potential and enhanced axonal anchoring. Methods: Genipin-crosslinked chitosan/PEG cryosponges were fabricated via focused graded functionalization of methoxy-PEG (mPEG) and blending with chitosan to obtain chitosan-Wend-mPEG (CbPEG-OH), chitosan-Wend-mPEG-CHO (CbPEG-CHO), chitosan-Wend-mPEG-NH_2 (CbPEG-NH_2) and chitosan-bfend-mPEG-COOH (CbPEG-COOH). The scaffolds were developed via a unique concurrent blending-crosslinking-cryogelling technique followed by incubation at -20°C and finally thawing at room temperature. Results and discussion: Morphologically, 7.5mm diameter cylindrical CbPEG-OH, CbPEG-CHO, CbPEG-NH_2 and CbPEG-COOH cryosponges composed of macroporous (discontinuous), uniaxial macroporous (continuous), mesoporous (continuous) and microporous (continuous) channels, respectively, of varying surface areas. The matrix resilience of the hydrated samples (at 25% strain) ranged between 12.7-17.5% on a force-time scale. The mechanical properties of CbPEGs prepared were within the range of bkjmectianical matrix rigidity of the rat spinal tissue tested (nanotensile testing). The neuronal anchoring of PC12 cells on the scaffolds was deduced via a relationship between the inherent matrix resilience (R_m; %), surface area (Ap) and solution viscosity (q) of the SpinoMimetic CryoSponges. At constant base/conjugate polymer ratio: A_p-√η/R_m(Equation 1) A_p =p√η/R_m (Equation 2) where, p (m~4s~4/kg) is Mechano-Mesoporous Flow Coefficient (novel biomaterial coefficient) which is a function of end group functionality of the conjugate polymer and expresses the proliferation and diffusion behaviour of PC12 cells dependent on local cytomechanical signals. The mathematical propositions in terms of p explained the spatiotemporal evolution of the growth factor concentration and neuronal connectivity (CbPEG-CHO ＞ CbPEG-OH ＞ CbPEG-NH_2 ＞ CbPEG-COOH). The neuronal anchoring performance of various cryosponges along with the energy relationships employing static lattice atomistic simulations (AMBER force field) demonstrated a significant role of torsional contributions arising from deviations from optimum dihedral angles as a function of degree of crosslinking. Conclusion: A novel relationship is hereby introduced for the prediction of behavior of axonal regeneration in response to substrate resilience, mechanical stress and topographic features of scaffolds with far reaching implications in spinal cord injury intervention.