3D Bioprinted Scaffold Remodels The Neuromodulatory Microenvironment For Enhancing Bone Regeneration

The bone tissue is densely populated with blood vessels and nerves with active metabolic activity.In recent years,several strategies have been developed to achieve vascularization in bone tissue engineering,but the communication between bone and nerves is not fully understood.Bone tissue is innervated by a large number of signals generated by sensory nerve fibers and sympathetic nerve fibers.Sensory nerves are located mainly in areas of active bone metabolism and are directly involved in osteogenesis through the secretion of neuropeptides such as calcitonin gene-related peptide(CGRP).However,studies have shown that activation of the sympathetic nervous system(SNS)may be detrimental to bone remodeling.Therefore,the pharmacological blockade of sympathetic signals could be a potential method for the treatment of bone defects.The 3D bioprinting technology has been used extensively in tissue engineering due to its ability to manufacture functional scaffolds with precise structures reproducibly,in which high-density living cells,biomaterials,and growth factors can be co-printed.Herein,we proposed a 3D bioprinted scaffold containing CGRP and the β-adrenergic receptor blocker propranolol(PRN)to achieve effective repair of bone defects.CGRP and PRN loaded mesoporous silica nanoparticles are added into a hybrid bio-ink,which initially contains gelatin methacrylate,Poly(ethylene glycol)diacrylate and bone marrow mesenchymal stem cells(BMSCs).Subsequently,3D bioprinting were used to create a composite scaffold with a pre-designed micro-nano hierarchical structure.In vitro studies showed that the mechanical,swelling,and degradation properties of Gel MA hydrogels were significantly improved with the incorporation of PEGDA.Meanwhile,the microporous structure inside the composite scaffold provided a favorable microenvironment for nutrient transport and cell attachment,proliferation,and migration.In vitro proliferation assays showed that the CGRP/PM/GP scaffold has good cytocompatibility,which could support cell proliferation.Drug release assays in vitro revealed that CGRP,PRN,and silica ions were sustained released from the scaffolds.With the release of CGRP from the scaffold,the secretion of neuropeptides by sensory nerves is simulated.The migration and tube formation of human umbilical vein endothelial cells(HUVECs)could be promoted by the CGRP/PM/GP scaffold,which is beneficial to the formation of a new capillary network during the bone repair process.In this study,the release of PRN can inhibit the binding process of catecholamine to β-adrenergic receptor,copromoting the osteogenic differentiation of BMSCs with CGRP and silicon ions,which will effectively enhance bone repairing of a critical-sized cranial defect in a rat model.In conclusion,this study provides a new idea for exploiting neuromodulation mechanisms during bone regeneration by constructing 3D bioprinting scaffolds that can remodel the neuromodulation microenvironment.