TrkA + sensory neurons regulate osteosarcoma proliferation and vascularization to promote disease progression.

Bone pain is a presenting feature of bone cancers such as osteosarcoma (OS), relayed by skeletal-innervating peripheral afferent neurons. Potential functions of tumor-associated sensory neurons in bone cancers beyond pain sensation are unknown. To uncover neural regulatory functions, a chemical-genetic approach in mice with a knock-in allele for TrkA was used to functionally perturb sensory nerve innervation during OS growth and disease progression. TrkA inhibition in transgenic mice led to significant reductions in sarcoma-associated sensory innervation and vascularization, tumor growth and metastasis, and prolonged overall survival. Single-cell transcriptomics revealed that sarcoma denervation was associated with phenotypic alterations in both OS tumor cells and cells within the tumor microenvironment, and with reduced calcitonin gene-related peptide (CGRP) and vascular endothelial growth factor (VEGF) signaling. Multimodal and multi-omics analyses of human OS bone samples and human dorsal root ganglia neurons further implicated peripheral innervation and neurotrophin signaling in OS tumor biology. In order to curb tumor-associated axonal ingrowth, we next leveraged FDA-approved bupivacaine liposomes leading to significant reductions in sarcoma growth, vascularity, as well as alleviation of pain. In sum, TrkA-expressing peripheral neurons positively regulate key aspects of OS progression and sensory neural inhibition appears to disrupt calcitonin receptor signaling (CALCR) and VEGF signaling within the sarcoma microenvironment leading to significantly reduced tumor growth and improved survival. These data suggest that interventions to prevent pathological innervation of osteosarcoma represent a novel adjunctive therapy to improve clinical outcomes and survival.

Mapping Somatosensory Afferent Circuitry to Bone Identifies Neurotrophic Signals Required for Fracture Healing.

The profound pain accompanying bone fracture is mediated by somatosensory neurons, which also appear to be required to initiate bone regeneration following fracture. Surprisingly, the precise neuroanatomical circuitry mediating skeletal nociception and regeneration remains incompletely understood. Here, we characterized somatosensory dorsal root ganglia (DRG) afferent neurons innervating murine long bones before and after experimental long bone fracture in mice. Retrograde labeling of DRG neurons by an adeno-associated virus with peripheral nerve tropism showed AAV-tdT signal. Single cell transcriptomic profiling of 6,648 DRG neurons showed highest labeling across CGRP+ neuron clusters (6.9-17.2%) belonging to unmyelinated C fibers, thinly myelinated Aδ fibers and Aß-Field LTMR (9.2%). Gene expression profiles of retrograde labeled DRG neurons over multiple timepoints following experimental stress fracture revealed dynamic changes in gene expression corresponding to the acute inflammatory ( S100a8 , S100a9 ) and mechanical force ( Piezo2 ). Reparative phase after fracture included morphogens such as Tgfb1, Fgf9 and Fgf18 . Two methods to surgically or genetically denervate fractured bones were used in combination with scRNA-seq to implicate defective mesenchymal cell proliferation and osteodifferentiation as underlying the poor bone repair capacity in the presence of attenuated innervation. Finally, multi-tissue scRNA-seq and interactome analyses implicated neuron-derived FGF9 as a potent regulator of fracture repair, a finding compatible with in vitro assessments of neuron-to-skeletal mesenchyme interactions.

Vegfc-expressing cells form heterotopic bone after musculoskeletal injury.

Heterotopic ossification (HO) is a challenging condition that occurs after musculoskeletal injury and is characterized by the formation of bone in non-skeletal tissues. While the effect of HO on blood vessels is well established, little is known about its impact on lymphatic vessels. Here, we use a mouse model of traumatic HO to investigate the relationship between HO and lymphatic vessels. We show that injury triggers lymphangiogenesis at the injury site, which is associated with elevated vascular endothelial growth factor C (VEGF-C) levels. Through single-cell transcriptomic analyses, we identify mesenchymal progenitor cells and tenocytes as sources of Vegfc. We demonstrate by lineage tracing that Vegfc-expressing cells undergo osteochondral differentiation and contribute to the formation of HO. Last, we show that Vegfc haploinsufficiency results in a nearly 50% reduction in lymphangiogenesis and HO formation. These findings shed light on the complex mechanisms underlying HO formation and its impact on lymphatic vessels.

Isolation of Perivascular Mesenchymal Progenitor Cells from Human Adipose Tissue by Flow Cytometry.

Perivascular cells represent an in vivo counterpart of mesenchymal stromal/stem cells that populate the outer layer of blood vessels. Pericytes in capillaries and microvessels and adventitial cells of large arteries and veins give rise to stem/progenitor cells when isolated and cultured in vitro. These cells have been considered candidate cell types for cell therapy. Adipose tissue, being highly vascularized, dispensable, and easily accessed, is a viable option to obtain perivascular cells for use in research and in clinical trials. Here, we describe our established protocol to extract perivascular cells from human fat through fluorescence-activated cell sorting, which allows for the isolation of defined populations of progenitor cells with high reproducibility.

Nr4a1 enhances Wnt4 transcription to promote mesenchymal stem cell osteogenesis and alleviates inflammation-inhibited bone regeneration.

Intense inflammatory response impairs bone marrow mesenchymal stem cell (BMSC)-mediated bone regeneration, with transforming growth factor (TGF)-ß1 being the most highly expressed cytokine. However, how to find effective and safe means to improve bone formation impaired by excessive TGF-ß1 remains unclear. In this study, we found that the expression of orphan nuclear receptor Nr4a1, an endogenous repressor of TGF-ß1, was suppressed directly by TGF-ß1-induced Smad3 and indirectly by Hdac4, respectively. Importantly, Nr4a1 overexpression promoted BMSC osteogenesis and reversed TGF-ß1-mediated osteogenic inhibition and pro-fibrotic effects. Transcriptomic and histologic analyses confirmed that upregulation of Nr4a1 increased the transcription of Wnt family member 4 (Wnt4) and activated Wnt pathway. Mechanistically, Nr4a1 bound to the promoter of Wnt4 and regulated its expression, thereby enhancing the osteogenic capacity of BMSCs. Moreover, treatment with Nr4a1 gene therapy or Nr4a1 agonist Csn-B could promote ectopic bone formation, defect repair, and fracture healing. Finally, we demonstrated the correlation of NR4A1 with osteogenesis and the activation of the WNT4/ß-catenin pathway in human BMSCs and fracture samples. Taken together, these findings uncover the critical role of Nr4a1 in bone formation and alleviation of inflammation-induced bone regeneration disorders, and suggest that Nr4a1 has the potential to be a therapeutic target for accelerating bone healing.

The use of weakly supervised machine learning for necrosis assessment in patients with osteosarcoma: A pilot study.

Percent necrosis (PN) following chemotherapy is a prognostic factor for survival in osteosarcoma. Pathologists estimate PN by calculating tumor viability over an average of whole-slide images (WSIs). This non-standardized, labor-intensive process requires specialized training and has high interobserver variability. Therefore, we aimed to develop a machine-learning model capable of calculating PN in osteosarcoma with similar accuracy to that of a musculoskeletal pathologist. In this proof-of-concept study, we retrospectively obtained six WSIs from two patients with conventional osteosarcomas. A weakly supervised learning model was trained by using coarse and incomplete annotations of viable tumor, necrotic tumor, and nontumor tissue in WSIs. Weakly supervised learning refers to processes capable of creating predictive models on the basis of partially and imprecisely annotated data. Once "trained," the model segmented areas of tissue and determined PN of the same six WSIs. To assess model fidelity, the pathologist also estimated PN of each WSI, and we compared the estimates using Pearson's correlation and mean absolute error (MAE). MAE was 15% over the six samples, and 6.4% when an outlier was removed, for which the model inaccurately labeled cartilaginous tissue. The model and pathologist estimates were strongly, positively correlated (r = 0.85). Thus, we created and trained a weakly supervised machine learning model to segment viable tumor, necrotic tumor, and nontumor and to calculate PN with accuracy similar to that of a musculoskeletal pathologist. We expect improvement can be achieved by annotating cartilaginous and other mesenchymal tissue for better representation of the histological heterogeneity in osteosarcoma.

TrkA-mediated sensory innervation of injured mouse tendon supports tendon sheath progenitor cell expansion and tendon repair.

Peripheral neurons terminate at the surface of tendons partly to relay nociceptive pain signals; however, the role of peripheral nerves in tendon injury and repair remains unclear. Here, we show that after Achilles tendon injury in mice, there is new nerve growth near tendon cells that express nerve growth factor (NGF). Conditional deletion of the Ngf gene in either myeloid or mesenchymal mouse cells limited both innervation and tendon repair. Similarly, inhibition of the NGF receptor tropomyosin receptor kinase A (TrkA) abrogated tendon healing in mouse tendon injury. Sural nerve transection blocked the postinjury increase in tendon sensory innervation and the expansion of tendon sheath progenitor cells (TSPCs) expressing tubulin polymerization promoting protein family member 3. Single cell and spatial transcriptomics revealed that disruption of sensory innervation resulted in dysregulated inflammatory signaling and transforming growth factor-ß (TGFß) signaling in injured mouse tendon. Culture of mouse TSPCs with conditioned medium from dorsal root ganglia neuron further supported a role for neuronal mediators and TGFß signaling in TSPC proliferation. Transcriptomic and histologic analyses of injured human tendon biopsy samples supported a role for innervation and TGFß signaling in human tendon regeneration. Last, treating mice after tendon injury systemically with a small-molecule partial agonist of TrkA increased neurovascular response, TGFß signaling, TSPC expansion, and tendon tissue repair. Although further studies should investigate the potential effects of denervation on mechanical loading of tendon, our results suggest that peripheral innervation is critical for the regenerative response after acute tendon injury.

Differential pericyte marker expression in craniofacial benign and malignant vascular tumors.

BACKGROUND: Vascular anomalies and tumors are common in the head, neck, and craniofacial areas and are associated with abnormalities in the angiomatous architecture. However, the etiology and molecular basis for the pathogenesis of most vascular lesions are still unknown. Pericytes are mural cells that surround endothelial cells. Besides angiogenesis and other physiological functions, pericytes play an important role in vascularized tissue repair and as resident mesenchymal stem/progenitor cells. Perivascular cells demonstrate a distinct immunohistochemical profile, including expression of alpha-smooth muscle actin (α-SMA), CD146, CD105, and PDGFRß, without endothelial differentiation (absence of CD31 and CD34 immunoreactivity). These pericyte markers have been shown to be expressed in soft tissue hemangiomas. However, they have not been fully examined in intraosseous hemangiomas. METHODS: In this study, we compared mesenchymal stem cell (MSC) expression of CD146 and α-SMA markers in pericytes from hemangiomas from different tissues and malignant vascular tumors. RESULTS: The results demonstrated an increased expression of pericyte markers in perivascular cells of benign hemangiomas, especially intraosseous hemangiomas and a significantly reduced expression of pericyte markers in malignant angiosarcomas. CONCLUSION: The evidence provides insight into the function of pericytes in vascular tumors and suggests their role in vascular tumor disease types.

ZIC1 Dictates Osteogenesis Versus Adipogenesis in Human Mesenchymal Progenitor Cells Via a Hedgehog Dependent Mechanism.

Numerous intrinsic factors regulate mesenchymal progenitor commitment to a specific cell fate, such as osteogenic or adipogenic lineages. Identification and modulation of novel intrinsic regulatory factors represent an opportunity to harness the regenerative potential of mesenchymal progenitors. In the present study, the transcription factor (TF) ZIC1 was identified to be differentially expressed among adipose compared with skeletal-derived mesenchymal progenitor cells. We observed that ZIC1 overexpression in human mesenchymal progenitors promotes osteogenesis and prevents adipogenesis. ZIC1 knockdown demonstrated the converse effects on cell differentiation. ZIC1 misexpression was associated with altered Hedgehog signaling, and the Hedgehog antagonist cyclopamine reversed the osteo/adipogenic differentiation alterations associated with ZIC1 overexpression. Finally, human mesenchymal progenitor cells with or without ZIC1 overexpression were implanted in an ossicle assay in NOD-SCID gamma mice. ZIC1 overexpression led to significantly increased ossicle formation in comparison to the control, as assessed by radiographic and histologic measures. Together, these data suggest that ZIC1 represents a TF at the center of osteo/adipogenic cell fate determinations-findings that have relevance in the fields of stem cell biology and therapeutic regenerative medicine.

Aldehyde Dehydrogenase, a Marker of Normal and Malignant Stem Cells, Typifies Mesenchymal Progenitors in Perivascular Niches.

Innate mesenchymal stem cells exhibiting multilineage differentiation and tissue (re)generative-or pathogenic-properties reside in perivascular niches. Subsets of these progenitors are committed to either osteo-, adipo-, or fibrogenesis, suggesting the existence of a developmental organization in blood vessel walls. We evaluated herein the activity of aldehyde dehydrogenase, a family of enzymes catalyzing the oxidation of aldehydes into carboxylic acids and a reported biomarker of normal and malignant stem cells, within human adipose tissue perivascular areas. A progression of ALDHLow to ALDHHigh CD34+ cells was identified in the tunica adventitia. Mesenchymal stem cell potential was confined to ALDHHigh cells, as assessed by proliferation and multilineage differentiation in vitro of cells sorted by flow cytometry with a fluorescent ALDH substrate. RNA sequencing confirmed and validated that ALDHHigh cells have a progenitor cell phenotype and provided evidence that the main isoform in this fraction is ALDH1A1, which was confirmed by immunohistochemistry. This demonstrates that ALDH activity, which marks hematopoietic progenitors and stem cells in diverse malignant tumors, also typifies native, blood vessel resident mesenchymal stem cells.

TIAM1 acts as an actin organization regulator to control adipose tissue-derived pericyte cell fate.

Pericytes are multipotent mesenchymal precursor cells that demonstrate tissue-specific properties. In this study, by comparing human adipose tissue- and periosteum-derived pericyte microarrays, we identified T cell lymphoma invasion and metastasis 1 (TIAM1) as a key regulator of cell morphology and differentiation decisions. TIAM1 represented a tissue-specific determinant between predispositions for adipocytic versus osteoblastic differentiation in human adipose tissue-derived pericytes. TIAM1 overexpression promoted an adipogenic phenotype, whereas its downregulation amplified osteogenic differentiation. These results were replicated in vivo, in which TIAM1 misexpression altered bone or adipose tissue generation in an intramuscular xenograft animal model. Changes in pericyte differentiation potential induced by TIAM1 misexpression correlated with actin organization and altered cytoskeletal morphology. Small molecule inhibitors of either small GTPase Rac1 or RhoA/ROCK signaling reversed TIAM1-induced morphology and differentiation in pericytes. In summary, our results demonstrate that TIAM1 regulates the cellular morphology and differentiation potential of human pericytes, representing a molecular switch between osteogenic and adipogenic cell fates.

CNTNAP4 signaling regulates osteosarcoma disease progression.

Improved treatment strategies for sarcoma rely on clarification of the molecular mediators of disease progression. Recently, we reported that the secreted glycoprotein NELL-1 modulates osteosarcoma (OS) disease progression in part via altering the sarcomatous extracellular matrix (ECM) and cell-ECM interactions. Of known NELL-1 interactor proteins, Contactin-associated protein-like 4 (Cntnap4) encodes a member of the neurexin superfamily of transmembrane molecules best known for its presynaptic functions in the central nervous system. Here, CRISPR/Cas9 gene deletion of CNTNAP4 reduced OS tumor growth, sarcoma-associated angiogenesis, and pulmonary metastases. CNTNAP4 knockout (KO) in OS tumor cells largely phenocopied the effects of NELL-1 KO, including reductions in sarcoma cell attachment, migration, and invasion. Further, CNTNAP4 KO cells were found to be unresponsive to the effects of NELL-1 treatment. Transcriptomic analysis combined with protein phospho-array demonstrated notable reductions in the MAPK/ERK signaling cascade with CNTNAP4 deletion, and the ERK1/2 agonist isoproterenol restored cell functions among CNTNAP4 KO tumor cells. Finally, human primary cells and tissues in combination with sequencing datasets confirmed the significance of CNTNAP4 signaling in human sarcomas. In summary, our findings demonstrate the biological importance of NELL-1/CNTNAP4 signaling axis in disease progression of human sarcomas and suggest that targeting the NELL-1/CNTNAP4 signaling pathway represents a strategy with potential therapeutic benefit in sarcoma patients.

Conventional Chondrosarcoma with Clear Cell Features in the Rib: Report of Two Cases and Review of the Literature.

A subset of clear cell chondrosarcomas may contain focal areas of low-grade conventional chondrosarcoma; however, it is rare to find foci resembling clear cell chondrosarcoma admixed with areas otherwise typical conventional chondrosarcoma. We report two patients with conventional chondrosarcoma with clear cell features occurring in the rib, one in the setting of multiple hereditary exostoses (MHE) and the other without MHE. Both patients were found to have a destructive rib mass with a soft tissue component and underwent en bloc resection. Histologic examination revealed predominantly grade 2 conventional chondrosarcomas; however, multiple foci containing large cells with pale eosinophilic to clear cytoplasm, distinct cell borders, centrally located nuclei, and conspicuous nucleoli, resembling clear cell chondrosarcoma were identified throughout the specimen. The significance of clear cell features in an otherwise typical conventional chondrosarcoma, to our knowledge, is unknown and deserves recognition. Finally, these tumors highlight the need for careful histologic examination and proper classification as unexpected findings may impact management.

Neurovascular coupling in bone regeneration.

The mammalian skeletal system is densely innervated by both neural and vascular networks. Peripheral nerves in the skeleton include sensory and sympathetic nerves. The crosstalk between skeletal and neural tissues is critical for skeletal development and regeneration. The cellular processes of osteogenesis and angiogenesis are coupled in both physiological and pathophysiological contexts. The cellular and molecular regulation of osteogenesis and angiogenesis have yet to be fully defined. This review will provide a detailed characterization of the regulatory role of nerves and blood vessels during bone regeneration. Furthermore, given the importance of the spatial relationship between nerves and blood vessels in bone, we discuss neurovascular coupling during physiological and pathological bone formation. A better understanding of the interactions between nerves and blood vessels will inform future novel therapeutic neural and vascular targeting for clinical bone repair and regeneration.

Label Cleaning Multiple Instance Learning: Refining Coarse Annotations on Single Whole-Slide Images.

Annotating cancerous regions in whole-slide images (WSIs) of pathology samples plays a critical role in clinical diagnosis, biomedical research, and machine learning algorithms development. However, generating exhaustive and accurate annotations is labor-intensive, challenging, and costly. Drawing only coarse and approximate annotations is a much easier task, less costly, and it alleviates pathologists' workload. In this paper, we study the problem of refining these approximate annotations in digital pathology to obtain more accurate ones. Some previous works have explored obtaining machine learning models from these inaccurate annotations, but few of them tackle the refinement problem where the mislabeled regions should be explicitly identified and corrected, and all of them require a - often very large - number of training samples. We present a method, named Label Cleaning Multiple Instance Learning (LC-MIL), to refine coarse annotations on a single WSI without the need for external training data. Patches cropped from a WSI with inaccurate labels are processed jointly within a multiple instance learning framework, mitigating their impact on the predictive model and refining the segmentation. Our experiments on a heterogeneous WSI set with breast cancer lymph node metastasis, liver cancer, and colorectal cancer samples show that LC-MIL significantly refines the coarse annotations, outperforming state-of-the-art alternatives, even while learning from a single slide. Moreover, we demonstrate how real annotations drawn by pathologists can be efficiently refined and improved by the proposed approach. All these results demonstrate that LC-MIL is a promising, lightweight tool to provide fine-grained annotations from coarsely annotated pathology sets.

Pharmacological inhibition of DKK1 promotes spine fusion in an ovariectomized rat model.

Osteoporosis is common in patients undergoing spine surgery, and carries a considerable risk of adverse outcomes. New methods to positively influence bone regeneration and spine fusion under osteoporotic conditions would be impactful. Neutralizing anti-Dickkopf-1 (DKK1) antibodies has been used as a bone anabolic agent, and recently reported by our group to aid in stem cell-mediated appendicular bone regeneration. Here, a small molecule designed as a DKK1 inhibitor, WAY-262611, was used to induce posterolateral spine fusion in an ovariectomized rat model. In vitro, pharmacological inhibition of DKK1 enhanced osteogenesis and Wnt signaling activity among rat bone marrow-derived stem/stromal cells (BMSCs). In vivo, systemic treatment with WAY-262611 promoted both chondrogenesis and osteogenesis within the spinal fusion site, and ultimately led to significant improvements in lumbar fusion as assessed by XR, µCT, histology and manual palpation assessments. No significant effect on osteoclast numbers or fusion site angiogenesis was detected, suggesting a primary direct effect on mesenchymal cells of the implantation site. Finally, evidence from human stem/stromal cells further demonstrated that pharmacologic inhibition of DKK1 promoted osteogenic differentiation in vitro. Taken together, our results suggest that targeting DKK1 promotes local bone formation and suggests potential clinical value for osteoporotic bone repair.

Acetabular Reaming Is a Reliable Model to Produce and Characterize Periarticular Heterotopic Ossification of the Hip.

Heterotopic ossification (HO) is a pathologic process characterized by the formation of bone tissue in extraskeletal locations. The hip is a common location of HO, especially as a complication of arthroplasty. Here, we devise a first-of-its-kind mouse model of post-surgical hip HO and validate expected cell sources of HO using several HO progenitor cell reporter lines. To induce HO, an anterolateral surgical approach to the hip was used, followed by disclocation and acetabular reaming. Animals were analyzed with high-resolution roentgenograms and micro-computed tomography, conventional histology, immunohistochemistry, and assessments of fluorescent reporter activity. All the treated animals' developed periarticular HO with an anatomical distribution similar to human patients after arthroplasty. Heterotopic bone was found in periosteal, inter/intramuscular, and intracapsular locations. Further, the use of either PDGFRα or scleraxis (Scx) reporter mice demonstrated that both cell types gave rise to periarticular HO in this model. In summary, acetabular reaming reproducibly induces periarticular HO in the mouse reproducing human disease, and with defined mesenchymal cellular contributors similar to other experimental HO models. This protocol may be used in the future for further detailing of the cellular and molecular mediators of post-surgical HO, as well as the screening of new therapies.

NELL1 Regulates the Matrisome to Promote Osteosarcoma Progression.

Sarcomas produce an abnormal extracellular matrix (ECM), which in turn provides instructive cues for cell growth and invasion. Neural EGF like-like molecule 1 (NELL1) is a secreted glycoprotein characterized by its nonneoplastic osteoinductive effects, yet it is highly expressed in skeletal sarcomas. Here, we show that genetic deletion of NELL1 markedly reduces invasive behavior across human osteosarcoma (OS) cell lines. NELL1 deletion resulted in reduced OS disease progression, inhibiting metastasis and improving survival in a xenograft mouse model. These observations were recapitulated with Nell1 conditional knockout in mouse models of p53/Rb-driven sarcomagenesis, which reduced tumor frequency and extended tumor-free survival. Transcriptomic and phosphoproteomic analyses demonstrated that NELL1 loss skews the expression of matricellular proteins associated with reduced FAK signaling. Culturing NELL1 knockout sarcoma cells on wild-type OS-enriched matricellular proteins reversed the phenotypic and signaling changes induced by NELL1 deficiency. In sarcoma patients, high expression of NELL1 correlated with decreased overall survival. These findings in mouse and human models suggest that NELL1 expression alters the sarcoma ECM, thereby modulating cellular invasive potential and prognosis. Disruption of NELL1 signaling may represent a novel therapeutic approach to short-circuit sarcoma disease progression. SIGNIFICANCE: NELL1 modulates the sarcoma matrisome to promote tumor growth, invasion, and metastasis, identifying the matrix-associated protein as an orchestrator of cell-ECM interactions in sarcomagenesis and disease progression.

Blood Vessel Resident Human Stem Cells in Health and Disease.

The vascular wall is comprised of distinct layers controlling angiogenesis, blood flow, vessel anchorage within organs, and cell and molecule transit between blood and tissues. Moreover, some blood vessels are home to essential stem-like cells, a classic example being the existence in the embryo of hemogenic endothelial cells at the origin of definitive hematopoiesis. In recent years, microvascular pericytes and adventitial perivascular cells were observed to include multi-lineage progenitor cells involved not only in organ turnover and regeneration but also in pathologic remodeling, including fibrosis and atherosclerosis. These perivascular mesodermal elements were identified as native forerunners of mesenchymal stem cells. We have presented in this brief review our current knowledge on vessel wall-associated tissue remodeling cells with respect to discriminating phenotypes, functional diversity in health and disease, and potential therapeutic interest.

NGF-p75 signaling coordinates skeletal cell migration during bone repair.

Bone regeneration following injury is initiated by inflammatory signals and occurs in association with infiltration by sensory nerve fibers. Together, these events are believed to coordinate angiogenesis and tissue reprogramming, but the mechanism of coupling immune signals to reinnervation and osteogenesis is unknown. Here, we found that nerve growth factor (NGF) is expressed following cranial bone injury and signals via p75 in resident mesenchymal osteogenic precursors to affect their migration into the damaged tissue. Mice lacking Ngf in myeloid cells demonstrated reduced migration of osteogenic precursors to the injury site with consequently delayed bone healing. These features were phenocopied by mice lacking p75 in Pdgfra+ osteoblast precursors. Single-cell transcriptomics identified mesenchymal subpopulations with potential roles in cell migration and immune response, altered in the context of p75 deletion. Together, these results identify the role of p75 signaling pathway in coordinating skeletal cell migration during early bone repair.