Neuronal Regulation of Skeletal Development and Repair

摘要 这是一项研究感觉神经在骨骼中的作用的程序的更新应用。我们的研究 在第一个资助期内证明了神经生长因子依赖的TrkA信号通过感觉神经是 在发育中的股骨和颅骨中血管生成和成骨的主要驱动力。在这些无血管的组织中 背景，间充质系细胞域中NGF的急性上调紧随其后的是伤害性纤维 向内生长，随后是间充质细胞增殖的部位。感觉受阻 神经生长，无论是通过抑制TrkA信号或破坏NGF，都会延缓血管形成和 破坏股骨和颅骨的骨形成。头盖骨模型中的组织学数据显示 感觉神经纤维的减少与增殖的间充质祖细胞数量减少有关 (MPC)在缝合和提前缝合。这些观察结果提出了一种范式，在这种范式中 感觉神经在骨骼发育中的作用，以维持MPC的可塑性，这一概念在 肢体再生模型，并得到最近小鼠股骨发育研究的支持。我们的预赛 在微流室中直接检测感觉神经轴突与基质细胞相互作用的研究结果 提示渗透的DRG神经纤维诱导MPC增殖，但限制非接触性MPC的分化 依附时尚。这些效应似乎是由神经来源的FSTL1介导的，FSTL1诱导MPC 增殖并损害BMP诱导的成骨分化。总而言之，这些数据支持这样一个前提 TrkA+感觉神经在骨骼发育过程中发挥作用，以维持干细胞的增殖， 通过递送激活有丝分裂和反分化的可溶性因子而处于未分化状态 信号通路。 这一概念模型将在分为两个具体目标的研究中进行探索。具体目标1将定义 发育中颅骨中TrkA+骨骼感觉神经的时空模式 对MPC增殖和细胞命运的影响，并进一步阐明与之相关的信号通路 神经受损。特定目标2将确定调节MPC增殖的感觉神经轴突衍生因子 和细胞命运决定，并最终确定FSTL1是影响MPC的神经源性因子 细胞行为。我们的结果应该会为感觉神经在脑内的基本作用提供新的见解 骨骼形态发生、动态平衡和修复，并提供对神经病理学的重要见解 与人类骨病相关的表现。

ABSTRACT This is a renewal application of a program investigating the role of sensory nerves in bone. Our studies during the first funding period demonstrate that NGF-dependent TrkA signaling by sensory nerves is the primary driver of angiogenesis and osteogenesis in the developing femur and skull. In these avascular settings, acute up-regulation of NGF in mesenchymal lineage cell domains is followed by nociceptive fiber ingrowth, which subsequently home to locations of proliferating mesenchymal cells. Blockade of sensory nerve ingrowth, either by inhibition of TrkA signaling or disruption of NGF, retards vascularization and disrupts femoral and calvarial bone formation. Histological data in the calvaria model revealed that loss of sensory nerve fibers is associated with reduced numbers of proliferating mesenchymal progenitor cells (MPCs) in the sutures and premature suture closure. These observations suggest a paradigm in which sensory nerves function in developing bone to maintain MPC plasticity, a concept well established in models of limb regeneration and supported by recent studies in developing mouse femur. Our preliminary findings directly examining the interaction of sensory nerve axons with MPCs in microfluidic chambers suggest that infiltrating DRG nerve fibers induce MPC proliferation, but limit differentiation in a non-contact dependent fashion. These effects appear to be mediated by neural derived FSTL1, which induces MPC proliferation and impairs BMP-induced osteogenic differentiation. Together, this data support the premise that TrkA+ sensory nerves function in developing bone to maintain stem cells in a proliferative, undifferentiated state by delivering soluble factors that activate mitogenic and anti-differentiation signaling pathways. This conceptual model will be explored in studies divided into two Specific Aims. Specific Aim 1 will define the spatiotemporal patterning of TrkA+ skeletal sensory nerves in the developing cranium, determine their influence on MPC proliferation and cellular fate, and further elucidate signaling pathways associated with impaired innervation. Specific Aim 2 will identify sensory axon-derived factors that regulate MPC proliferation and cell fate decisions, and definitively identifying FSTL1 as a neural-derived factor which impacts MPC cellular behavior. Our results should provide new insights into the fundamental roles sensory nerves play in skeletal morphogenesis, homeostasis and repair, and provide critical insight into the neuropathological manifestations associated with bone disorders in humans.