Heterotopic ossification (HO) is a debilitating condition characterized by the pathologic formation of ectopic bone. HO occurs commonly following orthopedic surgeries, burns, and neurologic injuries. ...While surgical excision may provide palliation, the procedure is often burdened with significant intra-operative blood loss due to a more robust contribution of blood supply to the pathologic bone than to native bone. Based on these clinical observations, we set out to examine the role of vascular signaling in HO. Vascular endothelial growth factor A (VEGFA) has previously been shown to be a crucial pro-angiogenic and pro-osteogenic cue during normal bone development and homeostasis. Our findings, using a validated mouse model of HO, demonstrate that HO lesions are highly vascular, and that VEGFA is critical to ectopic bone formation, despite lacking a contribution of endothelial cells within the developing anlagen.
Transforming growth factor-β1 (TGF-β1) plays a central role in normal and aberrant wound healing, but the precise mechanism in the local environment remains elusive. Here, using a mouse model of ...aberrant wound healing resulting in heterotopic ossification (HO) after traumatic injury, we find autocrine TGF-β1 signaling in macrophages, and not mesenchymal stem/progenitor cells, is critical in HO formation. In-depth single-cell transcriptomic and epigenomic analyses in combination with immunostaining of cells from the injury site demonstrated increased TGF-β1 signaling in early infiltrating macrophages, with open chromatin regions in TGF-β1-stimulated genes at binding sites specific for transcription factors of activated TGF-β1 (SMAD2/3). Genetic deletion of TGF-β1 receptor type 1 (Tgfbr1; Alk5), in macrophages, resulted in increased HO, with a trend toward decreased tendinous HO. To bypass the effect seen by altering the receptor, we administered a systemic treatment with TGF-β1/3 ligand trap TGF-βRII-Fc, which resulted in decreased HO formation and a delay in macrophage infiltration to the injury site. Overall, our data support the role of the TGF-β1/ALK5 signaling pathway in HO.
Inflammation after trauma is both critical to normal wound healing and may be highly detrimental when prolonged or unchecked with the potential to impair physiologic healing and promote
pathology. ...Mechanical strain after trauma is associated with impaired wound healing and increased inflammation. The exact mechanisms behind this are not fully elucidated. Neutrophil extracellular traps (NETs), a component of the neutrophil response to trauma, are implicated in a range of pro-inflammatory conditions. In the current study, we evaluated their role in linking movement and inflammation. We found that a link exists between the disruption and amplification of NETs which harbors the potential to regulate the wound's response to mechanical strain, while leaving the initial inflammatory signal necessary for physiologic wound healing intact.
Extracellular matrix (ECM) interactions regulate both the cell transcriptome and proteome, thereby determining cell fate. Traumatic heterotopic ossification (HO) is a disorder characterized by ...aberrant mesenchymal lineage (MLin) cell differentiation, forming bone within soft tissues of the musculoskeletal system following traumatic injury. Recent work has shown that HO is influenced by ECM-MLin cell receptor signaling, but how ECM binding affects cellular outcomes remains unclear. Using time course transcriptomic and proteomic analyses, we identified discoidin domain receptor 2 (DDR2), a cell surface receptor for fibrillar collagen, as a key MLin cell regulator in HO formation. Inhibition of DDR2 signaling, through either constitutive or conditional
deletion or pharmaceutical inhibition, reduced HO formation in mice. Mechanistically, DDR2 perturbation alters focal adhesion orientation and subsequent matrix organization, modulating Focal Adhesion Kinase (FAK) and Yes1 Associated Transcriptional Regulator and WW Domain Containing Transcription Regulator 1 (YAP/TAZ)-mediated MLin cell signaling. Hence, ECM-DDR2 interactions are critical in driving HO and could serve as a previously unknown therapeutic target for treating this disease process.
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