NLRC5在间充质干细胞增殖、归巢、成骨分化中的作用及机制研究

牙槽骨内源性修复及再生是组织再生的重要途径，而阐明免疫对成骨的调控机制是其重要前提。我们前期发现模式识别受体NLRC5影响小鼠的骨发育。在此基础上，本研究拟探索NLRC5在骨髓间充质干细胞（BMMSCs）增殖、归巢、成骨分化中的作用及机制。比较NLRC5-/-及野生型小鼠的mBMMSCs以及NLRC5-/-mBMMSCs表型逆转前后细胞增殖、凋亡及相关调控基因表达的差异；建立这两种小鼠荧光标记的BMMSCs移植合并牙槽骨缺损模型，比较两组mBMMSCs在骨缺损处归巢及归巢调控分子表达的差异；检测矿化诱导中成骨基因的表达及涉及的信号通路，并观察NLRC5对牙槽骨修复的影响；再通过改变人源性BMMSCs的NLRC5表达水平行验证实验，从BMMSCs增殖、归巢和成骨分化三方面阐述NLRC5对骨形成及修复的影响机制，进而从新的角度揭示骨修复再生的免疫调控机理，为临床应用提供理论基础及干预靶点。

Endogenous repair and regeneration of alveolar bone is a potential strategy for tissue regeneration, in which elucidation of the immunoregulatory mechanism of osteogenesis is indispensable. Our previous study discovered that NLRC5 might affect mice osteogenesis, and its mechanism deserves further research. Based on this, the present program aims to explore the role and mechanism of NLRC5 in proliferation, homing and osteogenesis of bone marrow mesenchymal stem cells. The detailed methods are as follows. Bone marrow derived mesenchymal stem cells （BMMSCs）from NLRC5 knockout (NLRC5-/-) and wild type (WT) mice and NLRC5-/- BMMSCs rescued by NLRC5 overexpression would be collected. Their proliferation, apoptosis as well as the activation of relative signal pathway would be detected. Recipient mice with alveolar bone defect would receive transplantation of NLRC5-/- or NLRC5+ BMMSCs carrying EGFP. BMMSCs homing in defected alveolar bone area would be detected by fluorescent microscope, and the expression of homing and co-receptor molecules would also be detected. In order to detect the role of NLRC5 in osteogenesis, the expression levels of osteogenic genes and relative signal pathway would be examined in NLRC5-/-and WT BMMSCs and in NLRC5-/- BMMSCs rescued by overexpression of NLRC5, and the alveolar bone restoration in recipient mice would also be investigated. The expression level of NLRC5 would be changed in human BMMSCs. Afterward, their proliferation, apoptosis, cycling regulating genes expression and the activation of relative signal pathway would be examined, in order to verify the involved mechanism. This program will supply evidences for the immune regulatory mechanism of bone regeneration from a new perspective, which sets up theoretical basis for a new treatment strategy of bone regeneration.