Release of O-GlcNAc transferase inhibitor promotes neuronal differentiation of neural stem cells in 3D bioprinted supramolecular hydrogel scaffold for spinal cord injury repair

Precise fabrication of biomimetic three-dimensional (3D) structure and effective neuronal differentiation under the pathological environment are the key to neural stem cell (NSC)-based spinal cord injury (SCI) therapy. In this study, we have developed a spinal cord-like bioprinted scaffold loading with OSMI-4, a small molecule O-GlcNAc transferase (OGT) inhibitor, to induce and guide the neuron differentiation of NSCs for efficient SCI repair. To achieve this, we developed a supramolecular bioink (SM bioink) consisting of methacrylated gelatin and acrylated β-cyclodextrins to load NSCs and OSMI-4. This bioink showed fast gelation and stable mechanical properties, facilitating bioprinting of functional neural scaffolds. Moreover, the weak host-guest cross-linking of the SM scaffolds significantly improved the cell-matrix interaction for the infiltration and migration of NSCs. What's more, the sustained delivery of OSMI-4 remarkably enhanced the intrinsic neuronal differentiation of the encapsulated NSCs in vitro by inhibiting Notch signaling pathway. In vivo experiment further revealed that the functional bioprinted scaffolds promoted the neuronal regeneration and axonal growth, leading to significant locomotor recovery of the SCI model rats. Together, the NSC-laden bioprinted SM scaffolds in combination with sustained release of the therapeutic agent OSMI-4 largely induced neuronal differentiation of NSCs and thus leading to efficient SCI repair. STATEMENT OF SIGNIFICANCE: Efficient neuronal differentiation of neural stem cells (NSCs) under the complex pathological microenvironment of spinal cord injury (SCI) is a major challenge of neural regeneration. By the use of a supramolecular bioink, we bioprinted a spinal cord-like scaffold loaded with NSCs and a small molecule drug OSMI-4 to significantly induce neuronal differentiation of NSCs for efficient SCI repair in vivo. The scaffolds with spinal cord-like structure can support the interaction and neuronal differentiation of NSCs by providing a dynamic matrix and a source of molecular release of OSMI-4. The influences of OSMI-4 on NSCs and its molecular mechanism were investigated for the first time in this study. Altogether, three-dimensional bioprinting fabrication of NSC- and small molecule drug-laden biomimetic construct may represent a promising therapeutic strategy for SCI repair.