组蛋白乙酰转移酶GCN5调控人间充质干细胞成骨向分化的作用和机制研究

Human mesenchymal stem cells (hMSCs) is one of the ideal seed cells for bone tissue engineering. As a histone acetyltransferase (HAT), GCN5 (general control nonderepressible 5) plays an essential role in the epigenetic regulation and growth development, however, little has been known about the osteogenesis of hMSCs. The study of our research group indicated that GCN5 can promote hMSCs osteogenesis, but the mechanism inside remains mysterious, thus inhibiting the protential transformation into clinical practice. NF-κB, a critical signaling pathway, has been found that played an important role in the osteogenesis of stem cells, but few studies were con-ducted about the upstream mechanism in these aspects..To explore and identify the whole epigenetic modulating pathway of “GCN5—NF-κB—osteoblastic differentiation of stem cells” can bring new insight for elevating the capacity of hMSCs osteogenesis epigenetically in the future. This project mainly includes the following aspects: explore the effects of GCN5 on the osteogenic differentiation of hMSCs (bone marrow, and adipose derived MSCs) and whether its HAT activity participates in the process; investigate the molecular mechanism between GCN5 and NF-κB, explore the modulation modes of GCN5 expression and functions of NF-κB and further effectively induce the osteoblastic differentiation of hMSCs; identify the osteogenic capacity of hMSCs when GCN5 involved as a drug target in vitro and in vivo. This project will establish a solid foundation for developing new target drugs which promote hMSCs osteogenesis, and make great benefits for the clinical transformation of stem cell technology.

人间充质干细胞（hMSCs）是骨组织工程理想的种子细胞之一。GCN5作为一种组蛋白乙酰转移酶，参与机体表观遗传学调控进程。尽管GCN5已被证实在生长发育中发挥重要作用，但是关于其诱导hMSCs成骨向分化的研究还未见报道。课题组前期研究发现GCN5可以促进hMSCs分化为成骨细胞，但其机制不明确。NF-κB作为细胞中关键的信号通路，参与干细胞成骨分化，但其上游调控机制的研究还很少。探索并明确“GCN5—NF-κB—干细胞成骨向分化”的整条表观遗传调控通路，可为将来在表观遗传学水平提高hMSCs成骨向分化能力提供一条新途径，对于干细胞技术的临床转化具有重大意义。项目内容：以人骨髓和脂肪MSCs为代表，研究GCN5表达水平对hMSCs成骨向分化的影响；探讨GCN5如何影响NF-κB通路，进而调控hMSCs的成骨向分化；在裸鼠体内和骨质疏松小鼠模型中检测GCN5调控hMSCs成骨向分化的能力。

该项目率先探索了H3K9乙酰化酶GCN5在间充质干细胞成骨分化过程中的关键调控作用。研究发现， GCN5对成骨分化的调控并不依赖其H3K9乙酰化酶活性。机制实验发现，GCN5通过抑制NF-kB信号通路来实现对间充质干细胞成骨分化的调控，这部分工作已发表于J Bone Min Res上；基于上述研究，我们还发现H3K9另外一个乙酰化酶PCAF也对成骨分化有重要调控作用。我们发现PCAF通过促进BMP信号通路相关基因启动子区H3K9乙酰化激活BMP通路，进而促进成骨分化，这部分工作发表于Stem Cells上；同时H3K9去乙酰化酶SIRT6也在成骨分化过程中发挥重要作用，但是SIRT6对成骨分化的调控并不依赖其H3K9去乙酰化酶活性，SIRT6协同PCAF调控BMP信号通路，促进间充质干细胞成骨分化，这部分工作发表于Sci Rep上。此外，在上述研究基础上，本项目还进一步深入研究了组蛋白H4R3甲基化酶PRMT3调控MSCs成骨分化的作用机制，这部分工作发表于Cell Death Dis上。本项目一方面为表观遗传调控成骨分化提供了新的理论依据，另一方面，也将有利于科学家们通过设计相应的小分子或化合物来改变干细胞染色质的结构或构型，为促进以MSCs为基础的骨组织工程或骨再生技术的临床转化奠定坚实基础。