Neuronal Regulation of Skeletal Development and Repair

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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 osteogenic precursors in the sutures and premature suture closure. These observations suggest a paradigm in which sensory nerves function in developing bone to maintain mesenchymal stem cell 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 MSCs in microfluidic chambers suggest that infiltrating DRG nerve fibers induce MSC proliferation, but limit differentiation in a non-contact dependent fashion. These effects are accompanied by upregulation of osteoprogenitor mitogens (e.g. TGF) and inhibitors of MSC differentiation (e.g. follistatin-like 1). 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 three Specific Aims. Specific Aim 1 will define the spatiotemporal patterning of TrkA+ skeletal sensory nerves in the developing cranium, and determine their influence on osteoprogenitor proliferation and cellular fate. Specific Aim 2 will identify key target genes in MSCs impacted by sensory nerve signals using previously validated co-culture methods. Specific Aim 3 will identify sensory axon-derived factors that regulate MSC proliferation and cell fate decisions. 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.