Cbfb基因在骨组织旁分泌通路调控骨髓脂肪组织形成中的功能研究

Osteoporosis is an extremely common age-linked condition that is characterized by low bone mass and decreased bone strength. Marrow adipose tissue (MAT) is usually considered to be deleterious to bone health since 1) increased marrow adipocyte formation is at the expense of osteoblast differentiation since both originate from marrow mesenchymal stem cell (MSC); 2) Paracrine factors secreted by MAT promotes bone resorption and inhibits bone formation. Given the importance of MAT in bone metabolism, molecular mechanism underlying MAT formation is still unclear, and paracrine factors within skeletal microenvironment that regulate MAT is also largely unknown. Deletion of Cbfβ in pre-osteoblast reduces bone mass and increases MAT in mice, partially through paracrine signaling. Thus, to characterize how MSC differentiation fate and MAT formation are regulated by osteocyte/mature osteoblast-secreted cytokines, we would like to generate osteoblast and osteocyte-specific Cbfb conditional knockout mouse models. We proposed to ①determine the role of osteocyte and osteoblast in MSC differentiation using a co-culture system; ②to characterize the role of Cbfb in late stage of osteoblast differentiation regarding bone metabolism and MAT; ③to characterize Cbfβ-regulated mature osteoblast/osteocyte-secreted cytokines that play an important role in MSC differentiation fate determination and MAT formation. The proposed study will lead to new understanding in molecular mechanism underlying MAT formation. The proposed study will also lead to discovery of novel pharmaceutical targets for osteoporosis treatment.

骨质疏松症是在老年人口中高发的、以骨量减少为特征的骨代谢疾病。由于骨髓间充质干细胞（MSC）成骨/成脂分化命运的“此消彼长”，以及骨髓脂肪组织（MAT）对骨代谢的抑制作用，MAT和骨量多呈现负相关的关系。目前MAT形成的分子机制尚不明确，骨骼微环境对MAT的调控亦缺少系统性研究，故解析MAT的形成机制具有重要的临床和科学意义。已知转录因子Cbfβ在成骨前体细胞中的缺陷导致小鼠骨量减少、MAT增加。本研究拟以成熟成骨细胞和骨细胞特异性Cbfβ敲除小鼠为模型，研究成骨细胞和骨细胞的旁分泌信号对MAT形成的调控机理。本研究拟①通过体外共培养实验确立成骨细胞和骨细胞对MSC分化方向的影响，②通过条件性敲除小鼠模型探讨成骨分化晚期Cbfβ的缺陷对代谢和MAT的影响，③通过高通量筛选寻找Cbfβ调控的、决定MSC分化和MAT形成的细胞因子。本研究将为骨质疏松症和MAT形成提供新的理论基础和靶点。

Cbfβ是Cbfb 基因编码的23KD 大小的转录调控因子，和RUNX 蛋白家族成员结合形成异二聚体CBF，特异性识别并结合共有结合DNA 序列，驱动目的基因的表达。CBFβ在早期成骨和间充质干细胞阶段促进成骨分化、增加骨量，功能较为明确；但其在成熟成骨细胞、骨细胞的功能还有待研究。本项目通过构建骨细胞、成熟成骨细胞Cbfb特异性敲除的小鼠模型，研究Cbfb在骨细胞中对骨代谢和骨微环境的调控作用，探索其背后的分子机制。我们的研究发现Cbfβ在骨细胞中对骨量起到负调控的作用，这与其已知的在间充质和成骨前体阶段的功能截然相反。通过组织学分析和RNAseq-转录组分析，我们发现Cbfβ调控了骨细胞的数量和小窝小管结构，同时控制骨细胞中分泌物（包括Tgfbeta1、Sfrp2、Sfrp4、OPG等）的表达，引起巨噬细胞活化和破骨细胞形成、调控骨吸收和骨稳态。同时Cbfβ在骨细胞中对骨脂的形成也起到抑制作用，但其对骨代谢的贡献不大。我们的研究解析了转录因子Cbfβ在骨细胞阶段全新的功能和机制，为骨细胞、骨脂在骨稳态和再生中的功能提供了重要的信息和思路。