A Mitochondrial-Based Transfer and Metabolic Remodeling Hydrogel with Immune Responsiveness to Augmented Bone Regeneration

Intercellular mitochondrial transfer has recently been discovered as a strategy for local microenvironment regulation and repair. However, improving the efficiency of mitochondrial transfer and reducing immune rejection caused by implants is challenging for its application in bone regeneration tissue engineering. We found that bone marrow mesenchymal stem cells (BMSC)s can obtain mitochondria from macrophages through tunnel nanotubes (TNT)s. Mitochondria are crucial for the metabolism and function of BMSC. We further designed a composite hydrogel based on aldehyde sodium hyaluronate and chitosan as the network matrix, containing anti-inflammatory dimethyl itaconate (DMI) and macrophage-targeted zwitterionic nanoparticles (MDVNPs). The acidic bone injury microenvironment triggers a reversible Schiff base reaction in response to pH; further, the released DMI upregulates the M2 phenotype of macrophages by reducing the ROS level. Subsequently, MDVNPs with targeting, high transfection, and low immunogenicity accelerated the efficiency of mitochondrial transfer between macrophages and BMSC by increasing the expression of the mitochondrial transfer regulatory protein Rho GTPase 1 (Miro1). In vitro studies have confirmed that promoting mitochondrial transfer can effectively increase adenosine triphosphate (ATP) and oxidative phosphorylation (OXPHOS) levels in BMSC, promoting osteoblastic differentiation. In the mouse model of critical defect, the composite hydrogel (Gel@MDI) can reduce inflammatory reactions, improve energy metabolism, and enhance bone repair by promoting the balance between the immune system and bone metabolism. This study establishes a potential method for the immune microenvironment to enhance cellular mitochondrial bioenergy and achieve tissue regeneration by regulating mitochondrial transfer between cells. We designed and developed immune responsiveness and injectable fluidity composite hydrogel (Gel@MDI) based on a reversible dynamic crosslinked network (-N=CH-), incorporating dimethyl itaconate (DMI) and macrophage-targeted zwitterionic nanoparticles (MDVNPs). Based on the Schiff-based reaction by aldehyde sodium hyaluronate (OSH) and chitosan (CS), hydrogel could realize the recombination and decomposition of polymer networks under different pH environments, which effectively induced the release of anti-inflammatory drug (DMI), leading to enhanced polarization of macrophages towards the M2 phenotype, thereby significantly improving the inflammatory response. Moreover, the novel zwitterionic nanoparticles (MDVNPs) not only effectively adjusted the mechanical properties of composite hydrogel to suit the bone regeneration microenvironment but also were capable of fulfilling their gene carrier function with exceptional efficiency, resulting in superior gene transfection rates and reduced cytotoxicity[19]. Therefore, Rho GTPase 1 (Miro1) loaded MDVNPs with macrophage-targeted function facilitated the mitochondrial transfer process from macrophages to BMSC. This transfer of mitochondria from macrophages serves as a crucial source of energy required for successful bone regeneration, which influences the intracellular metabolic state of BMSC by regulating ROS, adenosine triphosphate (ATP), and oxidative phosphorylation (OXPHOS) levels, thus promoting the formation and growth of fresh bone tissues[20]. Our study further explains the importance of bone immune regulation in the initiation stage of bone defect repair and the discovery of mitochondrial transfer as an intercellular regulatory approach in tissue regeneration, providing new ideas for tissue regeneration engineering.