Transient receptor potential vanilloid 1 interacts with Toll-like receptor 4 (TLR4)/cluster of differentiation 14 (CD14) signaling pathway in lipopolysaccharide-mediated inflammation in macrophages.

Transient receptor potential vanilloid 1 (TRPV1), a ligand-gated cation channel, is a receptor for vanilloids on sensory neurons and is also activated by capsaicin, heat, protons, arachidonic acid metabolites, and inflammatory mediators on neuronal or non-neuronal cells. However, the role of the TRPV1 receptor in pro-inflammatory cytokine secretion and its potential regulatory mechanisms in lipopolysaccharide (LPS)-induced inflammation has yet to be entirely understood. To investigate the role and regulatory mechanism of the TRPV1 receptor in regulating LPS-induced inflammatory responses, bone marrow-derived macrophages (BMDMs) harvested from wild-type (WT) and TRPV1 deficient (Trpv1-/-) mice were used as the cell model. In WT BMDMs, LPS induced an increase in the levels of tumor necrosis factor-α, IL-1ß, inducible nitric oxide synthase, and nitric oxide, which were attenuated in Trpv1-/- BMDMs. Additionally, the phosphorylation of inhibitor of nuclear factor kappa-Bα and mitogen-activated protein kinases, as well as the translocation of nuclear factor kappa-B and activator protein 1, were all decreased in LPS-treated Trpv1-/- BMDMs. Immunoprecipitation assay revealed that LPS treatment increased the formation of TRPV1-Toll-like receptor 4 (TLR4)-cluster of differentiation 14 (CD14) complex in WT BMDMs. Genetic deletion of TRPV1 in BMDMs impaired the LPS-triggered immune-complex formation of TLR4, myeloid differentiation protein 88, and interleukin-1 receptor-associated kinase, all of which are essential regulators in LPS-induced activation of the TLR4 signaling pathway. Moreover, genetic deletion of TRPV1 prevented the LPS-induced lethality and pro-inflammatory production in mice. In conclusion, the TRPV1 receptor may positively regulate the LPS-mediated inflammatory responses in macrophages by increasing the interaction with the TLR4-CD14 complex and activating the downstream signaling cascade.

Barriers and facilitators for recruiting and retaining male participants into longitudinal health research: a systematic review.

BACKGROUND: Successfully recruiting male participants to complete a healthcare related study is important for healthcare study completion and to advance our clinical knowledgebase. To date, most research studies have examined the barriers and facilitators of female participants in longitudinal healthcare-related studies with limited information available about the needs of males in longitudinal research. This systematic review examines the unique barriers and facilitators to male recruitment across longitudinal healthcare-related research studies. METHODS: Following PRIMSA guidelines, MEDLINE, Embase, CINAHL and Web of Science databases were systematically searched using the terms recruitment and/or retention, facilitators and/or barriers and longitudinal studies from 1900 to 2023 which contained separate data on males aged 17-59 years. Health studies or interventions were defined longitudinal if they were greater than or equal to 12 weeks in duration with 3 separate data collection visits. RESULTS: Twenty-four articles published from 1976-2023 met the criteria. One-third of the studies had a predominantly male sample and four studies recruited only male participants. Males appear disinterested towards participation in health research, however this lack of enthusiasm can be overcome by clear, non-directive communication, and studies that support the participants interests. Facilitating factors are diverse and may require substantial time from research teams. CONCLUSIONS: Future research should focus on the specific impact of these factors across the spectrum of longitudinal health-related studies. Based on the findings of this systematic review, researchers from longitudinal health-related clinical trials are encouraged to consider male-specific recruitment strategies to ensure successful recruitment and retention in their studies. REGISTRATION: This systemic review is registered with the PROSPERO database (CRD42021254696).

Bacterial Lipopolysaccharides Exacerbate Neurogenic Heterotopic Ossification Development.

Neurogenic heterotopic ossifications (NHO) are heterotopic bones that develop in periarticular muscles after severe central nervous system (CNS) injuries. Several retrospective studies have shown that NHO prevalence is higher in patients who suffer concomitant infections. However, it is unclear whether these infections directly contribute to NHO development or reflect the immunodepression observed in patients with CNS injury. Using our mouse model of NHO induced by spinal cord injury (SCI) between vertebrae T11 to T13 , we demonstrate that lipopolysaccharides (LPS) from gram-negative bacteria exacerbate NHO development in a toll-like receptor-4 (TLR4)-dependent manner, signaling through the TIR-domain-containing adapter-inducing interferon-ß (TRIF/TICAM1) adaptor rather than the myeloid differentiation primary response-88 (MYD88) adaptor. We find that T11 to T13 SCI did not significantly alter intestinal integrity nor cause intestinal bacteria translocation or endotoxemia, suggesting that NHO development is not driven by endotoxins from the gut in this model of SCI-induced NHO. Relevant to the human pathology, LPS increased expression of osteoblast markers in cultures of human fibro-adipogenic progenitors isolated from muscles surrounding NHO biopsies. In a case-control retrospective study in patients with traumatic brain injuries, infections with gram-negative Pseudomonas species were significantly associated with NHO development. Together these data suggest a functional association between gram-negative bacterial infections and NHO development and highlights infection management as a key consideration to avoid NHO development in patients. © 2023 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

Inflammasomes and the IL-1 Family in Bone Homeostasis and Disease.

PURPOSE OF REVIEW: Inflammasomes are multimeric protein structures with crucial roles in host responses against infections and injuries. The importance of inflammasome activation goes beyond host defense as a dysregulated inflammasome and subsequent secretion of IL-1 family members is believed to be involved in the pathogenesis of various diseases, some of which also produce skeletal manifestations. The purpose of this review is to summarize recent developments in the understanding of inflammasome regulation and IL-1 family members in bone physiology and pathology and current therapeutics will be discussed. RECENT FINDINGS: Small animal models have been vital to help understand how the inflammasome regulates bone dynamics. Animal models with gain or loss of function in various inflammasome components or IL-1 family signaling have illustrated how these systems can impact numerous bone pathologies and have been utilized to test new inflammasome therapeutics. It is increasingly clear that a tightly regulated inflammasome is required not only for host defense but for skeletal homeostasis, as a dysregulated inflammasome is linked to diseases of pathological bone accrual and loss. Given the complexities of inflammasome activation and redundancies in IL-1 activation and secretion, targeting these pathways is at times challenging. Ongoing research into inflammasome-mediated mechanisms will allow the development of new therapeutics for inflammasome/IL-1 diseases.

Lymphocytes Are Not Required for Neurogenic Heterotopic Ossification Development after Spinal Cord Injury.

Neurogenic heterotopic ossifications (NHOs) are incapacitating complications of traumatic brain and spinal cord injuries (SCI) that manifest as abnormal bone formation in periarticular muscles. Using a unique model of NHO after SCI in genetically unmodified mice, we have previously established that the innate immune system plays a key driving role in NHO pathogenesis. The role of adaptive immune cells in NHO pathogenesis, however, remains unexplored in this model. Here we established that B lymphocytes were reduced in the spleen and blood after SCI and increased in muscles of mice in which NHO develops, whereas minimal changes in T cell frequencies were noted. Interestingly, Rag1 -/- mice lacking mature T and B lymphocytes, developed NHO, similar to wild-type mice. Finally, mice that underwent splenectomy before SCI and muscle damage also developed NHO to the same extent as non-splenectomized SCI controls. Overall, our findings show that functional T and B lymphocytes have minimal influence or dispensable contributions to NHO development after experimental SCI in mice.

Spinal cord injury reprograms muscle fibroadipogenic progenitors to form heterotopic bones within muscles.

The cells of origin of neurogenic heterotopic ossifications (NHOs), which develop frequently in the periarticular muscles following spinal cord injuries (SCIs) and traumatic brain injuries, remain unclear because skeletal muscle harbors two progenitor cell populations: satellite cells (SCs), which are myogenic, and fibroadipogenic progenitors (FAPs), which are mesenchymal. Lineage-tracing experiments using the Cre recombinase/LoxP system were performed in two mouse strains with the fluorescent protein ZsGreen specifically expressed in either SCs or FAPs in skeletal muscles under the control of the Pax7 or Prrx1 gene promoter, respectively. These experiments demonstrate that following muscle injury, SCI causes the upregulation of PDGFRα expression on FAPs but not SCs and the failure of SCs to regenerate myofibers in the injured muscle, with reduced apoptosis and continued proliferation of muscle resident FAPs enabling their osteogenic differentiation into NHOs. No cells expressing ZsGreen under the Prrx1 promoter were detected in the blood after injury, suggesting that the cells of origin of NHOs are locally derived from the injured muscle. We validated these findings using human NHO biopsies. PDGFRα+ mesenchymal cells isolated from the muscle surrounding NHO biopsies could develop ectopic human bones when transplanted into immunocompromised mice, whereas CD56+ myogenic cells had a much lower potential. Therefore, NHO is a pathology of the injured muscle in which SCI reprograms FAPs to undergo uncontrolled proliferation and differentiation into osteoblasts.

Oncostatin M regulates hematopoietic stem cell (HSC) niches in the bone marrow to restrict HSC mobilization.

We show that pro-inflammatory oncostatin M (OSM) is an important regulator of hematopoietic stem cell (HSC) niches in the bone marrow (BM). Treatment of healthy humans and mice with granulocyte colony-stimulating factor (G-CSF) dramatically increases OSM release in blood and BM. Using mice null for the OSM receptor (OSMR) gene, we demonstrate that OSM provides a negative feed-back acting as a brake on HSPC mobilization in response to clinically relevant mobilizing molecules G-CSF and CXCR4 antagonist. Likewise, injection of a recombinant OSM molecular trap made of OSMR complex extracellular domains enhances HSC mobilization in poor mobilizing C57BL/6 and NOD.Cg-PrkdcscidIl2rgtm1Wjl/SzJ mice. Mechanistically, OSM attenuates HSC chemotactic response to CXCL12 and increases HSC homing to the BM signaling indirectly via BM endothelial and mesenchymal cells which are the only cells expressing OSMR in the BM. OSM up-regulates E-selectin expression on BM endothelial cells indirectly increasing HSC proliferation. RNA sequencing of HSCs from Osmr-/- and wild-type mice suggest that HSCs have altered cytoskeleton reorganization, energy usage and cycling in the absence of OSM signaling in niches. Therefore OSM is an important regulator of HSC niche function restraining HSC mobilization and anti-OSM therapy combined with current mobilizing regimens may improve HSPC mobilization for transplantation.

Interleukin-1 Is Overexpressed in Injured Muscles Following Spinal Cord Injury and Promotes Neurogenic Heterotopic Ossification.

Neurogenic heterotopic ossifications (NHOs) form in periarticular muscles after severe spinal cord (SCI) and traumatic brain injuries. The pathogenesis of NHO is poorly understood with no effective preventive treatment. The only curative treatment remains surgical resection of pathological NHOs. In a mouse model of SCI-induced NHO that involves a transection of the spinal cord combined with a muscle injury, a differential gene expression analysis revealed that genes involved in inflammation such as interleukin-1ß (IL-1ß) were overexpressed in muscles developing NHO. Using mice knocked-out for the gene encoding IL-1 receptor (IL1R1) and neutralizing antibodies for IL-1α and IL-1ß, we show that IL-1 signaling contributes to NHO development after SCI in mice. Interestingly, other proteins involved in inflammation that were also overexpressed in muscles developing NHO, such as colony-stimulating factor-1, tumor necrosis factor, or C-C chemokine ligand-2, did not promote NHO development. Finally, using NHO biopsies from SCI and TBI patients, we show that IL-1ß is expressed by CD68+ macrophages. IL-1α and IL-1ß produced by activated human monocytes promote calcium mineralization and RUNX2 expression in fibro-adipogenic progenitors isolated from muscles surrounding NHOs. Altogether, these data suggest that interleukin-1 promotes NHO development in both humans and mice. © 2021 American Society for Bone and Mineral Research (ASBMR).

Local and Systemic Factors Drive Ectopic Osteogenesis in Regenerating Muscles of Spinal-Cord-Injured Mice in a Lesion-Level-Dependent Manner.

Neuroimmune dysfunction is thought to promote the development of several acute and chronic complications in spinal cord injury (SCI) patients. Putative roles for adrenal stress hormones and catecholamines are increasingly being recognized, yet how these adversely affect peripheral tissue homeostasis and repair under SCI conditions remains elusive. Here, we investigated their influence in a mouse model of SCI with acquired neurogenic heterotopic ossification. We show that spinal cord lesions differentially influence muscular regeneration in a level-dependent manner and through a complex multi-step process that creates an osteopermissive environment within the first hours of injury. This cascade of events is shown to critically involve adrenergic signals and drive the acute release of the neuropeptide, substance P. Our findings generate new insights into the kinetics and processes that govern SCI-induced deregulations in skeletal muscle homeostasis and regeneration, thereby aiding the development of sequential therapeutic strategies that can prevent or attenuate neuromusculoskeletal complications in SCI patients.

Neurogenic Heterotopic Ossifications Recapitulate Hematopoietic Stem Cell Niche Development Within an Adult Osteogenic Muscle Environment.

Hematopoiesis and bone interact in various developmental and pathological processes. Neurogenic heterotopic ossifications (NHO) are the formation of ectopic hematopoietic bones in peri-articular muscles that develop following severe lesions of the central nervous system such as traumatic cerebral or spinal injuries or strokes. This review will focus on the hematopoietic facet of NHO. The characterization of NHO demonstrates the presence of hematopoietic marrow in which quiescent hematopoietic stem cells (HSC) are maintained by a functional stromal microenvironment, thus documenting that NHOs are neo-formed ectopic HSC niches. Similarly to adult bone marrow, the NHO permissive environment supports HSC maintenance, proliferation and differentiation through bidirectional signaling with mesenchymal stromal cells and endothelial cells, involving cell adhesion molecules, membrane-bound growth factors, hormones, and secreted matrix proteins. The participation of the nervous system, macrophages and inflammatory cytokines including oncostatin M and transforming growth factor (TGF)-ß in this process, reveals how neural circuitry fine-tunes the inflammatory response to generate hematopoietic bones in injured muscles. The localization of NHOs in the peri-articular muscle environment also suggests a role of muscle mesenchymal cells and bone metabolism in development of hematopoiesis in adults. Little is known about the establishment of bone marrow niches and the regulation of HSC cycling during fetal development. Similarities between NHO and development of fetal bones make NHOs an interesting model to study the establishment of bone marrow hematopoiesis during development. Conversely, identification of stage-specific factors that specify HSC developmental state during fetal bone development will give more mechanistic insights into NHO.

When the Nervous System Turns Skeletal Muscles into Bones: How to Solve the Conundrum of Neurogenic Heterotopic Ossification.

PURPOSE OF REVIEW: Neurogenic heterotopic ossification (NHO) is the abnormal formation of extra-skeletal bones in periarticular muscles after damage to the central nervous system (CNS) such as spinal cord injury (SCI), traumatic brain injury (TBI), stroke, or cerebral anoxia. The purpose of this review is to summarize recent developments in the understanding of NHO pathophysiology and pathogenesis. Recent animal models of NHO and recent findings investigating the communication between CNS injury, tissue inflammation, and upcoming NHO therapeutics are discussed. RECENT FINDINGS: Animal models of NHO following TBI or SCI have shown that NHO requires the combined effects of a severe CNS injury and soft tissue damage, in particular muscular inflammation and the infiltration of macrophages into damaged muscles plays a key role. In the context of a CNS injury, the inflammatory response to soft tissue damage is exaggerated and persistent with excessive signaling via substance P-, oncostatin M-, and TGF-ß1-mediated pathways. This review provides an overview of the known animal models and mechanisms of NHO and current therapeutic interventions for NHO patients. While some of the inflammatory mechanisms leading to NHO are common with other forms of traumatic and genetic heterotopic ossifications (HO), NHOs uniquely involve systemic changes in response to CNS injury. Future research into these CNS-mediated mechanisms is likely to reveal new targetable pathways to prevent NHO development in patients.

Neurogenic Heterotopic Ossifications Develop Independently of Granulocyte Colony-Stimulating Factor and Neutrophils.

Neurogenic heterotopic ossifications (NHOs) are incapacitating heterotopic bones in periarticular muscles that frequently develop following traumatic brain or spinal cord injuries (SCI). Using our unique model of SCI-induced NHO, we have previously established that mononucleated phagocytes infiltrating injured muscles are required to trigger NHO via the persistent release of the pro-inflammatory cytokine oncostatin M (OSM). Because neutrophils are also a major source of OSM, we investigated whether neutrophils also play a role in NHO development after SCI. We now show that surgery transiently increased granulocyte colony-stimulating factor (G-CSF) levels in blood of operated mice, and that G-CSF receptor mRNA is expressed in the hamstrings of mice developing NHO. However, mice defective for the G-CSF receptor gene Csf3r, which are neutropenic, have unaltered NHO development after SCI compared to C57BL/6 control mice. Because the administration of recombinant human G-CSF (rhG-CSF) has been trialed after SCI to increase neuroprotection and neuronal regeneration and has been shown to suppress osteoblast function at the endosteum of skeletal bones in human and mice, we investigated the impact of a 7-day rhG-CSF treatment on NHO development. rhG-CSF treatment significantly increased neutrophils in the blood, bone marrow, and injured muscles. However, there was no change in NHO development compared to saline-treated controls. Overall, our results establish that unlike monocytes/macrophages, neutrophils are dispensable for NHO development following SCI, and rhG-CSF treatment post-SCI does not impact NHO development. Therefore, G-CSF treatment to promote neuroregeneration is unlikely to adversely promote or affect NHO development in SCI patients. © 2020 American Society for Bone and Mineral Research.

Inhibition of JAK1/2 Tyrosine Kinases Reduces Neurogenic Heterotopic Ossification After Spinal Cord Injury.

Neurogenic heterotopic ossifications (NHO) are very incapacitating complications of traumatic brain and spinal cord injuries (SCI) which manifest as abnormal formation of bone tissue in periarticular muscles. NHO are debilitating as they cause pain, partial or total joint ankylosis and vascular and nerve compression. NHO pathogenesis is unknown and the only effective treatment remains surgical resection, however once resected, NHO can re-occur. To further understand NHO pathogenesis, we developed the first animal model of NHO following SCI in genetically unmodified mice, which mimics most clinical features of NHO in patients. We have previously shown that the combination of (1) a central nervous system lesion (SCI) and (2) muscular damage (via an intramuscular injection of cardiotoxin) is required for NHO development. Furthermore, macrophages within the injured muscle play a critical role in driving NHO pathogenesis. More recently we demonstrated that macrophage-derived oncostatin M (OSM) is a key mediator of both human and mouse NHO. We now report that inflammatory monocytes infiltrate the injured muscles of SCI mice developing NHO at significantly higher levels compared to mice without SCI. Muscle infiltrating monocytes and neutrophils expressed OSM whereas mouse muscle satellite and interstitial cell expressed the OSM receptor (OSMR). In vitro recombinant mouse OSM induced tyrosine phosphorylation of the transcription factor STAT3, a downstream target of OSMR:gp130 signaling in muscle progenitor cells. As STAT3 is tyrosine phosphorylated by JAK1/2 tyrosine kinases downstream of OSMR:gp130, we demonstrated that the JAK1/2 tyrosine kinase inhibitor ruxolitinib blocked OSM driven STAT3 tyrosine phosphorylation in mouse muscle progenitor cells. We further demonstrated in vivo that STAT3 tyrosine phosphorylation was not only significantly higher but persisted for a longer duration in injured muscles of SCI mice developing NHO compared to mice with muscle injury without SCI. Finally, administration of ruxolitinib for 7 days post-surgery significantly reduced STAT3 phosphorylation in injured muscles in vivo as well as NHO volume at all analyzed time-points up to 3 weeks post-surgery. Our results identify the JAK/STAT3 signaling pathway as a potential therapeutic target to reduce NHO development following SCI.

CD169+ macrophages are critical for osteoblast maintenance and promote intramembranous and endochondral ossification during bone repair.

Osteal macrophages (osteomacs) contribute to bone homeostasis and regeneration. To further distinguish their functions from osteoclasts, which share many markers and growth factor requirements, we developed a rapid, enzyme-free osteomac enrichment protocol that permitted characterization of minimally manipulated osteomacs by flow cytometry. Osteomacs differ from osteoclasts in expression of Siglec1 (CD169). This distinction was confirmed using the CD169-diphtheria toxin (DT) receptor (DTR) knock-in model. DT treatment of naïve CD169-DTR mice resulted in selective and striking loss of osteomacs, whilst osteoclasts and trabecular bone area were unaffected. Consistent with a previously-reported trophic interaction, osteomac loss was accompanied by a concomitant and proportionately striking reduction in osteoblasts. The impact of CD169+ macrophage depletion was assessed in two models of bone injury that heal via either intramembranous (tibial injury) or endochondral (internally-plated femoral fracture model) ossification. In both models, CD169+ macrophage, including osteomac depletion compromised bone repair. Importantly, DT treatment in CD169-DTR mice did not affect osteoclast frequency in either model. In the femoral fracture model, the magnitude of callus formation correlated with the number of F4/80+ macrophages that persisted within the callus. Overall these observations provide compelling support that CD169+ osteomacs, independent of osteoclasts, provide vital pro-anabolic support to osteoblasts during both bone homeostasis and repair.

Macrophage-derived oncostatin M contributes to human and mouse neurogenic heterotopic ossifications.

Neurogenic heterotopic ossification (NHO) is the formation of ectopic bone generally in muscles surrounding joints following spinal cord or brain injury. We investigated the mechanisms of NHO formation in 64 patients and a mouse model of spinal cord injury-induced NHO. We show that marrow from human NHOs contains hematopoietic stem cell (HSC) niches, in which mesenchymal stromal cells (MSCs) and endothelial cells provide an environment supporting HSC maintenance, proliferation, and differentiation. The transcriptomic signature of MSCs from NHOs shows a neuronal imprinting associated with a molecular network required for HSC support. We demonstrate that oncostatin M (OSM) produced by activated macrophages promotes osteoblastic differentiation and mineralization of human muscle-derived stromal cells surrounding NHOs. The key role of OSM was confirmed using an experimental model of NHO in mice defective for the OSM receptor (OSMR). Our results provide strong evidence that macrophages contribute to NHO formation through the osteogenic action of OSM on muscle cells within an inflammatory context and suggest that OSM/OSMR could be a suitable therapeutic target. Altogether, the evidence of HSCs in ectopic bones growing at the expense of soft tissue in spinal cord/brain-injured patients indicates that inflammation and muscle contribute to HSC regulation by the brain-bone-blood triad.

Characterization of Normal Murine Carpal Bone Development Prompts Re-Evaluation of Pathologic Osteolysis as the Cause of Human Carpal-Tarsal Osteolysis Disorders.

Multicentric carpal-tarsal osteolysis; multicentric osteolysis, nodulosis, and arthropathy; and Winchester syndromes, skeletal dysplasias characterized by carpal/tarsal and epiphyseal abnormalities, are caused by mutations in v-maf musculoaponeurotic fibrosarcoma oncogene ortholog B (MAFB), matrix metalloproteinase (MMP) 2, and MMP14, respectively; however, the underlying pathophysiology is unclear. Osteoclast-mediated osteolysis has been regarded as the main mechanism, but does not explain the skeletal distribution. We hypothesized that MAFB, MMP-2, and MMP-14 have integral roles in carpal/tarsal and epiphyseal bone development. Normal neonatal mouse forepaws were imaged by micro-computed tomography and examined histologically. Murine forepaw ossification occurred sequentially. Subarticular regions of endochondral ossification showed morphologic and calcification patterns that were distinct from archetypical physeal endochondral ossification. This suggests that two different forms of endochondral ossification occur. The skeletal sites showing the greatest abnormality in the carpal-tarsal osteolysis syndromes are regions of subarticular ossification. Thus, abnormal bone formation in areas of subarticular ossification may explain the site-specific distribution of the carpal-tarsal osteolysis phenotype. MafB, Mmp-2, and Mmp-14 were expressed widely, and tartrate-resistant acid phosphatase staining notably was absent in the subarticular regions of the cartilage anlagen and entheses at a time point most relevant to the human osteolysis syndromes. Thus, abnormal peri-articular skeletal development and modeling, rather than excessive bone resorption, may be the underlying pathophysiology of these skeletal syndromes.

Early anti-inflammatory intervention ameliorates axial disease in the proteoglycan-induced spondylitis mouse model of ankylosing spondylitis.

BACKGROUND: Ankylosing spondylitis (AS) is characterised by immune-mediated arthritis and osteoproliferation, ultimately leading to joint ankylosis. Whether inflammation is necessary for osteoproliferation is controversial, fuelled by the unclear efficacy of anti-inflammatory treatments on radiographic progression. In proteoglycan-induced spondylitis (PGISp), a mouse model of AS, inflammation is the prerequisite for osteoproliferation as osteoproliferation was only observed following inflammation-driven intervertebral disc (IVD) destruction. We hypothesised that early intervention with a potent anti-inflammatory therapy would protect IVD integrity and consequently alter disease progression. METHODS: PGISp mice received vehicle or a combination of etanercept (ETN) plus prednisolone (PRD) therapy for 2 or 6 weeks initiated at an early disease stage. Peripheral arthritis was scored longitudinally. Spinal disease was assessed using a semi-quantitative histological scoring regimen including inflammation, joint destruction and excessive tissue formation. RESULTS: ETN + PRD therapy significantly delayed the onset of peripheral arthritis. IVD integrity was significantly protected when treatment was commenced in early disease. Six-weeks of treatment resulted in trends towards reductions in intervertebral joint damage and excessive tissue formation. IVD score distribution was dichotomized, likely reflecting the extent of axial disease at initiation of therapy. In the sub-group of mice with high IVD destruction scores, ETN + PRD treatment significantly reduced IVD destruction severity, inflammation and bone erosion and reduced cartilage damage and excessive tissue formation. CONCLUSIONS: Early intervention with anti-inflammatory treatment not only improved inflammatory symptoms but also ameliorated structural damage of spine in PGISp mice. This preclinical observation suggests that early anti-inflammatory intervention may slow radiographic progression in AS patients.

CD169(+) macrophages mediate pathological formation of woven bone in skeletal lesions of prostate cancer.

Skeletal metastases present a major clinical challenge for prostate cancer patient care, inflicting distinctive mixed osteoblastic and osteolytic lesions that cause morbidity and refractory skeletal complications. Macrophages are abundant in bone and bone marrow and can influence both osteoblast and osteoclast function in physiology and pathology. Herein, we examined the role of macrophages in prostate cancer bone lesions, particularly the osteoblastic response. First, macrophage and lymphocyte distributions were qualitatively assessed in patient's prostate cancer skeletal lesions by immunohistochemistry. Second, macrophage functional contributions to prostate tumour growth in bone were explored using an immune-competent mouse model combined with two independent approaches to achieve in vivo macrophage depletion: liposome encapsulated clodronate that depletes phagocytic cells (including macrophages and osteoclasts); and targeted depletion of CD169(+) macrophages using a suicide gene knock-in model. Immunohistochemistry and histomorphometric analysis were performed to quantitatively assess cancer-induced bone changes. In human bone metastasis specimens, CD68(+) macrophages were consistently located within the tumour mass. Osteal macrophages (osteomacs) were associated with pathological woven bone within the metastatic lesions. In contrast, lymphocytes were inconsistently present in prostate cancer skeletal lesions and when detected, had varied distributions. In the immune-competent mouse model, CD169(+) macrophage ablation significantly inhibited prostate cancer-induced woven bone formation, suggesting that CD169(+) macrophages within pathological woven bone are integral to tumour-induced bone formation. In contrast, pan-phagocytic cell, but not targeted CD169(+) macrophage depletion resulted in increased tumour mass, indicating that CD169(-) macrophage subset(s) and/or osteoclasts influenced tumour growth. In summary, these observations indicate a prominent role for macrophages in prostate cancer bone metastasis that may be therapeutically targetable to reduce the negative skeletal impacts of this malignancy, including tumour-induced bone modelling. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.