The molecular regulation of NGF-mediated differentiation of dental pulp stem cells

项目总结/摘要 R 03提案的总体目标是研究NGF在牙髓中的促成骨作用 在体外和体内研究干细胞（DPSC），并应用这些知识促进DPSC在 多种临床环境。DPSC是多能祖细胞，其可容易地从提取的或培养的细胞中获得。 脱落的牙齿，并具有高矿化潜力和增殖率。因此，DPSC介导的 治疗在再生骨缺损方面取得了临床成功， 颅面疾病我们最近的工作是利用细胞表面活性剂分离DPSC的同质群体， 标志物表达显示，CD 271是分离高成骨能力的DPSC的最可靠的标志物。 潜力此外，用已知的CD 271配体NGF处理，促进了骨形成分化。 私营部门安保公司。当我们检查NGF的细胞内信号传导途径时，我们发现ALK和JNK是 在NGF介导的DPSC成骨过程中上调，证实了基于受体的信号转导。后 通过检查表观遗传调节因子，我们发现NGF显著诱导KDM 4 B的表达，KDM 4 B是一种组蛋白， 去甲基酶负责去除沉默标记，H3 K9 me 3。因此，我们假设， 通过CD 271信号传导和表观遗传调节增强DPSC的成骨分化， 成骨基因（例如，RUNX 2和DLX 5）。这一假设将通过以下具体目标进行检验：1） 确定CD 271受体的功能参与，并描绘其在NGF- 在体外和体内诱导DPSC的成骨分化; 2）阐明表观遗传的作用， 在NGF介导的DPSC分化中的调节剂。目前的提案有望揭示 神经生长因子介导的DPSC成骨机制的见解，对颅面 组织工程学联合DPSC和NGF治疗可能是未来的创新答案。 再生疗法不仅基于其上级成骨能力，而且基于其在神经发生中的作用 和血管生成，所有这些都为再生创造了有利的微环境。 总结：制定当前R 03提案是为了为后续R 01积累初步数据 应用程序. R 01的未来研究将结合联合收割机两个新兴领域，表观遗传学和组织工程学， 牙科MSC，在颅面缺损再生的临床工具的发展。其他角色 将探讨牙科MSC功能和分化的表观遗传学，阐明复杂的相互作用， 以及表观遗传和遗传之间的协同相互作用 因素将评估KDM 4 B敲除小鼠以确定KDM 4 B在颅面发育中的作用。 此外，使用R 03提案中的小鼠模型，KDM 4 B在骨组织工程中的作用也将 进行评估。

PROJECT SUMMARY/ABSTRACT The overall objectives of this R03 proposal are to investigate the pro-osteogenic effects of NGF in dental pulp stem cells (DPSCs) in vitro and in vivo and to apply this knowledge to foster the application of DPSCs in multiple clinical settings. DPSCs are multipotent progenitor cells that are readily available from extracted or exfoliated teeth and possess high mineralization potential and proliferation rates. As such, DPSC-mediated therapy has produced clinical success in regenerating bone defects and consequently holds great promise for craniofacial disorders. Our recent work in isolating homogenous populations of DPSCs using cell surface marker expression revealed that CD271 was the most reliable marker to isolate DPSCs with high osteogenic potential. Furthermore, treatment with NGF, a known ligand for CD271, promoted osteogenic differentiation of DPSCs. When we examined the intracellular signaling pathways of NGF, we found that ALK and JNK were upregulated during NGF-mediated DPSC osteogenesis, confirming receptor based signal transduction. Upon examining epigenetic regulators, we found that NGF significantly induced expression of KDM4B, a histone demethylase responsible for removing the silencing mark, H3K9me3. We therefore hypothesize that NGF enhances osteogenic differentiation of DPSCs through CD271 signaling and epigenetic modulation of osteogenic genes (e.g., RUNX2 and DLX5). This hypothesis will be tested by the following specific aims: 1) To determine the functional involvement of CD271 receptor and delineate its intracellular signaling in NGF- induced osteogenic differentiation in DPSCs both in vitro and in vivo; 2) To elucidate the role of epigenetic regulators in NGF-mediated DPSC differentiation. The current proposal holds the promise of revealing the mechanistic insights of NGF-mediated osteogenesis in DPSCs, with significant implications for craniofacial tissue engineering. Combined DPSC and NGF therapy may be the innovative answer for the future of regenerative therapy on the basis of not only its superior osteogenic capacity but also its role in neurogenesis and angiogenesis, all of which create a favorable microenvironment for regeneration. Summary: The current R03 proposal is formulated to accumulate preliminary data for subsequent R01 application. Future studies in R01 will combine two emerging areas, epigenetics and tissue engineering using dental MSCs, in the development of clinical tools in craniofacial defect regeneration. Additional roles of epigenetics in dental MSC function and differentiation will be explored, elucidating the complex interplay between various epigenetic modulators and the synergistic interactions between epigenetic and genetic factors. KDM4B knockout mice will be evaluated to determine the role of KDM4B in craniofacial development. Additionally, using the mouse model from this R03 proposal, KDM4B's role in bone tissue engineering will also be assessed.