Precise in-situ release of microRNA from an injectable hydrogel induces bone regeneration

Abstract Critical bone defects are a common yet challenging orthopedic problem. Tissue engineering is an emerging and promising strategy for bone regeneration in large-scale bone defects. The precise on-demand release of osteogenic factors is critical for controlling the osteogenic differentiation of seed cells with the support of appropriate three dimensional scaffolds. However, most of the effective osteogenic factors are biomacromolecules with release behaviors that are difficult to control. Here, the cholesterol-modified non-coding microRNA Chol-miR-26a was used to promote the osteogenic differentiation of human mesenchymal stem cells (hMSCs). Chol-miR-26a was conjugated to an injectable poly(ethylene glycol) (PEG) hydrogel through an ultraviolet (UV)-cleavable ester bond. The injectable PEG hydrogel was formed by a copper-free click reaction between the terminal azide groups of 8-armed PEG and dibenzocyclooctyne-biofunctionalized PEG, into which UV-cleavable Chol-miR-26a was simultaneously conjugated via a Michael addition reaction. Upon UV irradiation, Gel-c-miR-26a (MLCaged) released Chol-c-miR-26a selectively and exhibited significantly improved efficacy in bone regeneration compared to the hydrogel without UV irradiation and UV-uncleavable MLControl. MLCaged significantly enhanced alkaline phosphatase activity and promoted calcium nodule deposition in vitro and repaired critical skull defects in a rat animal model, demonstrating that injectable implantation with the precise release of osteogenic factors has the potential to repair large-scale bone defects in clinical practice. Statement of significance Provide a novel and practical strategy via hydrogel for efficient delivery and precisely controlled release of miRNAs into bone defect sites. The hydrogel is formed by polyethylene glycol (PEG), which is crosslinked by ‘click’ reaction. Cholesterol-modified miR-26a loading on the hydrogel is covalently patterned onto the fibers of hydrogel through a UV light-cleavable linker, which prevents undesired release of miRNA. This hydrogel could realize the controlled release of miRNA under light regulation both in vitro and in vivo, thus realize bone regeneration.