组蛋白甲基化修饰骨髓基质干细胞修复眼眶骨缺损的作用机制研究

The orbital walls are thin, and adjacent to paranasal sinuses with poor blood supply, where the defects are difficult to repair and may lead to visual dysfunction and facial deformities. Previous studies showed that osteogenic induction of bone marrow mesenchymal stem cell (BMSC) can heighten an effect on orbital bone defects repairing. Histone methylation in nucleosome structure can significantly enhance osteogenic differentiation and bone repair potential of BMSC, The H3K4 demethylase, Jumonji AT-rich interactive domain 1A (Jarid1a) is an important protein by regulating histone methylation level. This proposed study is to systematically investigate the function and regulatory mechanism of histone methylation in osteogenic differentiation of BMSC, and repair the canine medial orbital bone defects using histone methylated BMSC and β- tricalcium phosphate (β-TCP) scaffold. Our approach is to investigate that: ① The effect of gene modification by demethylase Jarid1a on osteogenic differentiation of BMSC; ② The regulatory mechanism of osteogenic gene expression by histone methylation; ③ Detecting in situ osteogenesis of the histone methylated BMSC composited with porous β-TCP scaffold by implantation in vivo, repairing the canine orbital bone defects and investigating the potentials of histone methylated BMSC in orbital bone regeneration. This study will systematically explore the function and mechanisms of histone methylated BMSC in orbital bone defect repair at the levels of gene, protein as well as small and large animals, and be expected to provide new approach for clinical reconstruction of orbital bone defects.

眼眶骨壁菲薄，毗邻副鼻窦空腔，局部血供差，创伤缺损后易造成患者视功能障碍与面部畸形。前期研究表明骨向诱导分化可增强骨髓基质干细胞（BMSC）修复眼眶骨缺损的能力，课题组发现脱甲基化酶Jarid1a可通过甲基化修饰组蛋白显著增强BMSC的成骨分化和骨修复能力。本项目将系统性研究组蛋白甲基化对BMSC成骨分化的调控作用机制和眼眶骨缺损的修复潜能。研究包括：①脱甲基酶Jarid1a基因修饰对BMSC成骨分化的作用；②组蛋白甲基化修饰对成骨基因表达的调控机制；③将组蛋白甲基化修饰BMSC复合磷酸三钙支架材料移植到体内，检测其原位促成骨作用，并用于犬眼眶内侧壁骨缺损模型的修复重建中，探讨其促眼眶骨再生作用。从基因-蛋白-小动物-大动物4个层次，多角度探索组蛋白甲基化在眼眶骨缺损修复中的作用和相关机制，有望为临床上眼眶骨缺损的修复重建提供新方法。

眼眶骨壁菲薄，毗邻副鼻窦空腔，局部血供差，创伤缺损后易造成患者视功能障碍与面部畸形。前期研究表明骨向诱导分化可增强骨髓基质干细胞（BMSC）修复眼眶骨缺损的能力，课题组发现脱甲基化酶Jarid1a可通过甲基化修饰组蛋白显著增强BMSC的成骨分化和骨修复潜能。本项目将系统性研究组蛋白甲基化修饰对BMSC的成骨分化作用及调控机制，进一步用于眼眶骨缺损的修复。研究包括：①脱甲基酶Jarid1a基因修饰对BMSC成骨分化的作用；②组蛋白甲基化修饰对成骨转录基因表达的调控机制；③将组蛋白甲基化修饰BMSC复合三磷酸钙支架材料移植到体内，检测其原位促成骨作用，并用于犬眼眶内侧壁骨缺损模型的修复重建中，探讨其促眼眶骨再生作用。我们科研工作完成了原定目标，通过大鼠BMSCs细胞的原代分离培养，构建Jarid1a过表达和抑制的慢病毒载体，并转染细胞，分子生物学及形态学方法显示转染抑制Jarid1a显著增强BMSCs的成骨分化水平,抑制Jarid1a显著促进BMSC 碱性磷酸酶活性和矿化作用，发现Jarid1a与Runx2的直接结合显著抑制BMSCs成骨分化。通过Jarid1a过表达和抑制的慢病毒载体转染大鼠BMSCs细胞，将细胞与支架材料复合后形成细胞材料复合体，修复大鼠颅骨标准骨缺损模型，组织切片等组织形态学方法评估显示抑制Jarid1a明显提高了BMSCs对骨缺损的修复效果。另外，我们还对眼眶新型材料及临床上眼眶骨折修复术进行了初期研究。相关成果在国内外学术刊物上发表标有本项目资助的论文 8篇，其中 SCI 论文 7篇，协助培养研究生4名，参加国内学术会议2次，大会发言1次。

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