Despite serving as the clinical “gold standard” treatment for critical size bone defects, decellularized allografts suffer from long-term failure rates of ~60% due to the absence of the periosteum. Stem and osteoprogenitor cells within the periosteum orchestrate autograft healing through host cell recruitment, which initiates the regenerative process. To emulate periosteum-mediated healing, tissue engineering approaches have been utilized with mixed outcomes. While vascularization has been widely established as critical for bone regeneration, innervation was recently identified to be spatiotemporally regulated together with vascularization and similarly indispensable to bone healing. Notwithstanding, there are no known approaches that have focused on periosteal matrix cues to coordinate host vessel and/or axon recruitment. Here, we investigated the influence of hydrogel degradation mechanism, i.e. hydrolytic or enzymatic (cell-dictated), on tissue engineered periosteum (TEP)-modified allograft healing, especially host vessel/nerve recruitment and integration. Matrix metalloproteinase (MMP)-degradable hydrogels supported endothelial cell migration from encapsulated spheroids whereas no migration was observed in hydrolytically degradable hydrogels in vitro, which correlated with increased neurovascularization in vivo. Specifically, ~2.45 and 1.84-fold, and ~3.48 and 2.58-fold greater vessel and nerve densities with high levels of vessel and nerve co-localization was observed using MMP degradable TEP (MMP-TEP) -modified allografts versus unmodified and hydrolytically degradable TEP (Hydro-TEP)-modified allografts, respectively, at 3 weeks post-surgery. MMP-TEP-modified allografts exhibited greater longitudinal graft-localized vascularization and endochondral ossification, along with 4-fold and 2-fold greater maximum torques versus unmodified and Hydro-TEP-modified allografts after 9 weeks, respectively, which was comparable to that of autografts. In summary, our results demonstrated that the MMP-TEP coordinated allograft healing via early stage recruitment and support of host neurovasculature.

尽管脱细胞同种异体移植是治疗临界尺寸骨缺损的临床“金标准”，但由于缺乏骨膜，其长期失败率约为 60%。骨膜内的干细胞和骨祖细胞通过宿主细胞募集协调自体移植物愈合，从而启动再生过程。为了模拟骨膜介导的愈合，组织工程方法已被采用，但结果好坏参半。虽然血管化已被广泛认为对骨再生至关重要，但最近发现神经支配与血管化一起受到时空调节，并且同样对于骨愈合不可或缺。尽管如此，还没有已知的方法专注于骨膜基质线索来协调宿主血管和/或轴突募集。在这里，我们研究了水凝胶降解机制，即水解或酶促（细胞决定）对组织工程骨膜（TEP）修饰的同种异体移植愈合的影响，特别是宿主血管/神经募集和整合。基质金属蛋白酶（MMP）可降解水凝胶支持内皮细胞从封装球体迁移，而在体外水解可降解水凝胶中未观察到迁移，这与体内神经血管化的增加相关。具体而言，使用 MMP 可降解 TEP (MMP-TEP) 修饰的同种异体移植物与未修饰和可水解降解的同种异体移植物相比，观察到血管和神经密度高约 2.45 倍和 1.84 倍、约 3.48 倍和 2.58 倍，且具有高水平的血管和神经共定位术后 3 周分别进行 TEP (Hydro-TEP) 修饰的同种异体移植物。 9周后，MMP-TEP修饰的同种异体移植物表现出更大的纵向移植物局部血管化和软骨内骨化，最大扭矩分别比未修饰和Hydro-TEP修饰的同种异体移植物高4倍和2倍，这与未修饰的同种异体移植物相当。自体移植物。总之，我们的结果表明，MMP-TEP 通过早期募集和支持宿主神经血管系统来协调同种异体移植愈合。