预诱导软骨分化促进颌骨再生的效应及机制研究

Maxillofacial bone defects or inflammatory stimuli often cause local environmental ischemia and hypoxia, leading to osteogenesis imperfecta or bone graft failure. Cell survival and the establishment of the blood supply has become the key to solve the problem. Due to its strong resistance to ischemia and hypoxia, chondrocytes has been suggested to be applied in maxillofacial bone defects, but the poor clinical outcome restricted its application, some researchers believed that cartilage transplantation does not apply to the membrane bone. Our group and other research group found that the hypertrophic cartilage derived from pre-induction treatment benefit in vivo bone formation, even in the membrane bone, while the mechanism is still unknown. Combined with the concept of development engineering, we proposed that pre-induction strategy of chondrogenic differentiation could be applied in maxillofacial defects, and the possible mechanisms to promote pre-differentiated cartilage into bone are: after implantation of pre-differentiated cartilage into the bone defect, cartilage could chemotactic mesenchymal stem cells, endothelial progenitor cell and other cells migrate to the defect site, interacts with each other, and ultimately improve bone formation; meanwhile hypertrophic chondrocytes in the cartilage block could differentiate into osteoblasts, then strengthen osteogenesis. Preliminary experiments showed that this approach could be the key to resolve the osteogenesis imperfecta or bone grafts failure coursed by critical ischemia and hypoxia, and provide new ideas and new methods for maxillofacial bone regeneration.

缺血缺氧局部环境是导致口腔颌面部骨缺损和种植体周骨缺损成骨不全或骨移植失败的重要原因。如何使细胞耐受恶劣环境并尽快建立血供是解决此难题的关键。由于软骨细胞对缺血缺氧环境有较强的抵抗力，多年前软骨块移植治疗颌骨缺损曾被寄予厚望，但因其效果不佳被否决。有学者认为软骨成骨的策略并不适用于膜内成骨来源的颌骨。但是，Martin组和本课题组都发现预诱导处理的肥大软骨细胞形成的组织工程软骨可在体内异位成骨，而且我们又发现它在膜内成骨来源的牙槽骨和颅骨内同样有效。这一现象背后的机制不明。因此，本项目提出预诱导软骨分化促进颌骨成骨的两个可能机制：肥大软骨细胞募集间充质干细胞、内皮祖细胞和单核巨噬细胞等迁移到缺损部位，并相互作用最终促进成骨；同时软骨块中的肥大软骨细胞向成骨细胞分化，进一步加强成骨。本课题建立在大量前期研究基础上，预诱导软骨块移植有望为颌骨和牙周骨再生、种植体周骨缺损治疗提供新思路和新方法。

缺血缺氧局部环境是导致口腔颌面部骨缺损和种植体周骨缺损、成骨不全或骨移植失败的重要原因。如何使细胞耐受恶劣环境并尽快建立血供是解决此难题的关键。由于软骨细胞对缺血缺氧环境有较强的抵抗力，多年前软骨块移植治疗颌骨缺损曾被寄予厚望。因此，本项目根据前期研究基础，提出预诱导软骨块作为一种新型的模拟体内软骨内成骨发育过程的组织工程媒介，能够有效地治疗各类骨缺损模型，甚至包括颌骨这类利用膜内成骨发育而来的骨。在本项目中，我们制备并优化了3D打印钛合金支架及羟基磷灰石涂层，载入骨髓来源间充质干细胞（bone mesenchymal stem cells，BMSCs）预诱导成软骨块；之后，利用大鼠下颌骨缺损模型，验证预诱导软骨块在修复颌骨缺损中的有效性；最后，证实预诱导软骨块在骨生成过程中与受体其他免疫细胞（尤其是巨噬细胞）间的相互作用、重要联系及相关机制；此外，我们还证实一种芳香烃受体（AhR）激活剂—FICZ防治牙周炎的效果及其相关机制。实验结果表明，相比于传统的利用BMSCs直接作为一种种子细胞治疗骨缺损的方法，预诱导软骨块体现出了更好的效果，一方面是体现在最终成骨效果更佳，另一方面，体外试验提示预诱导软骨块可能比未分化的BMSCs具有更良好的免疫调节能力，预诱导软骨块促巨噬细胞M2向极化而抑制其M1向极化，而两种表型的巨噬细胞都可以不同程度地促进软骨块成骨成血管进程。从骨免疫的角度来说，这具有重大的意义。因为任何植入物植入体内后，都先要与体内的各类免疫细胞发生作用，能够更好地调节局部免疫微环境的植入物，不仅自身具有更大的可能性存活下来，而且可以利用局部免疫微环境，帮助达到最后的成骨成血管的效果。这给临床上组织工程的利用巨大的启发。本课题建立在大量前期研究基础上，预诱导软骨块移植有望为颌骨和牙周骨再生、种植体周骨缺损治疗提供新思路和新方法。