Osteoimmunology of Fracture Healing.

PURPOSE OF REVIEW: The purpose of this review is to summarize what is known in the literature about the role inflammation plays during bone fracture healing. Bone fracture healing progresses through four distinct yet overlapping phases: formation of the hematoma, development of the cartilaginous callus, development of the bony callus, and finally remodeling of the fracture callus. Throughout this process, inflammation plays a critical role in robust bone fracture healing. RECENT FINDINGS: At the onset of injury, vessel and matrix disruption lead to the generation of an inflammatory response: inflammatory cells are recruited to the injury site where they differentiate, activate, and/or polarize to secrete cytokines for the purposes of cell signaling and cell recruitment. This process is altered by age and by sex. Bone fracture healing is heavily influenced by the presence of inflammatory cells and cytokines within the healing tissue.

Bone and Cytokine Markers Associated With Bone Disease in Systemic Mastocytosis.

BACKGROUND: Mastocytosis encompasses a heterogeneous group of diseases characterized by tissue accumulation of clonal mast cells, which frequently includes bone involvement. Several cytokines have been shown to play a role in the pathogenesis of bone mass loss in systemic mastocytosis (SM), but their role in SM-associated osteosclerosis remains unknown. OBJECTIVE: To investigate the potential association between cytokine and bone remodeling markers with bone disease in SM, aiming at identifying biomarker profiles associated with bone loss and/or osteosclerosis. METHODS: A total of 120 adult patients with SM, divided into 3 age and sex-matched groups according to their bone status were studied: (1) healthy bone (n = 46), (2) significant bone loss (n = 47), and (3) diffuse bone sclerosis (n = 27). Plasma levels of cytokines and serum baseline tryptase and bone turnover marker levels were measured at diagnosis. RESULTS: Bone loss was associated with significantly higher levels of serum baseline tryptase (P = .01), IFN-γ (P = .05), IL-1ß (P = .05), and IL-6 (P = .05) versus those found in patients with healthy bone. In contrast, patients with diffuse bone sclerosis showed significantly higher levels of serum baseline tryptase (P < .001), C-terminal telopeptide (P < .001), amino-terminal propeptide of type I procollagen (P < .001), osteocalcin (P < .001), bone alkaline phosphatase (P < .001), osteopontin (P < .01), and the C-C Motif Chemokine Ligand 5/RANTES chemokine (P = .01), together with lower IFN-γ (P = .03) and RANK-ligand (P = .04) plasma levels versus healthy bone cases. CONCLUSIONS: SM with bone mass loss is associated with a proinflammatory cytokine profile in plasma, whereas diffuse bone sclerosis shows increased serum/plasma levels of biomarkers related to bone formation and turnover, in association with an immunosuppressive cytokine secretion profile.

Considerations for understanding protein measurements: Identification of formation, degradation and more pathological relevant epitopes.

OBJECTIVES: There is a need for precision medicine and an unspoken promise of an optimal approach for identification of the right patients for value-based medicine based on big data. However, there may be a misconception that measurement of proteins is more valuable than measurement of fewer selected biomarkers. In population-based research, variation may be somewhat eliminated by quantity. However, this fascination of numbers may limit the attention to and understanding of the single. This review highlights that protein measurements (with collagens as examples) may mean different things depending on the targeted epitope - formation or degradation of tissues, and even signaling potential of proteins. DESIGN AND METHODS: PubMed was searched for collagen, neo-epitope, biomarkers. RESULTS: Ample examples of assays with specific epitopes, either pathological such as HbA1c, or domain specific such as pro-peptides, which total protein arrays would not have identified were evident. CONCLUSIONS: We suggest that big data may be considered as the funnel of data points, in which most important parameters will be selected. If the technical precision is low or the biological accuracy is limited, and we include suboptimal quality of biomarkers, disguised as big data, we may not be able to fulfill the promise of helping patients searching for the optimal treatment. Alternatively, if the technical precision of the total protein quantification is high, but we miss the functional domains with the most considerable biological meaning, we miss the most important and valuable information of a given protein. This review highlights that measurements of the same protein in different ways may provide completely different meanings. We need to understand the pathological importance of each epitope quantified to maximize protein measurements.

The Macrophage-Osteoclast Axis in Osteoimmunity and Osteo-Related Diseases.

Osteoimmunity is involved in regulating the balance of bone remodeling and resorption, and is essential for maintaining normal bone morphology. The interaction between immune cells and osteoclasts in the bone marrow or joint cavity is the basis of osteoimmunity, in which the macrophage-osteoclast axis plays a vital role. Monocytes or tissue-specific macrophages (macrophages resident in tissues) are an important origin of osteoclasts in inflammatory and immune environment. Although there are many reports on macrophages and osteoclasts, there is still a lack of systematic reviews on the macrophage-osteoclast axis in osteoimmunity. Elucidating the role of the macrophage-osteoclast axis in osteoimmunity is of great significance for the research or treatment of bone damage caused by inflammation and immune diseases. In this article, we introduced in detail the concept of osteoimmunity and the mechanism and regulators of the differentiation of macrophages into osteoclasts. Furthermore, we described the role of the macrophage-osteoclast axis in typical bone damage caused by inflammation and immune diseases. These provide a clear knowledge framework for studying macrophages and osteoclasts in inflammatory and immune environments. And targeting the macrophage-osteoclast axis may be an effective strategy to treat bone damage caused by inflammation and immune diseases.

The Roles of FOXO1 in Periodontal Homeostasis and Disease.

Periodontitis is an oral chronic inflammatory disease that is initiated by periodontal microbial communities and requires disruption of the homeostatic responses. The prevalence of periodontal disease increases with age; more than 70% of adults 65 years and older have periodontal disease. A pathogenic microbial community is required for initiating periodontal disease. Dysbiotic immune-inflammatory response and bone remodeling are characteristics of periodontitis. The transcription factor forkhead box protein O1 (FOXO1) is a key regulator of a number of cellular processes, including cell survival and differentiation, immune status, reactive oxygen species (ROS) scavenging, and apoptosis. Although accumulating evidence indicates that FOXO1 activity can be induced by periodontal pathogens, the roles of FOXO1 in periodontal homeostasis and disease have not been well documented. The present review summarizes how the FOXO1 signaling axis can regulate periodontal bacteria-epithelial interactions, immune-inflammatory response, bone remodeling, and wound healing.

Potential role of myeloid-derived suppressor cells in transition from reaction to repair phase of bone healing process.

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of immature myeloid cells with immunosuppressive functions; these cells play a key role in infection, immunization, chronic inflammation, and cancer. Recent studies have reported that immunosuppression plays an important role in the healing process of tissues and that Treg play an important role in fracture healing. MDSCs suppress active T cell proliferation and reduce the severity of arthritis in mice and humans. Together, these findings suggest that MDSCs play a role in bone biotransformation. In the present study, we examined the role of MDSCs in the bone healing process by creating a bone injury at the tibial epiphysis in mice. MDSCs were identified by CD11b and GR1 immunohistochemistry and their role in new bone formation was observed by detection of Runx2 and osteocalcin expression. Significant numbers of MDSCs were observed in transitional areas from the reactionary to repair stages. Interestingly, MDSCs exhibited Runx2 and osteocalcin expression in the transitional area but not in the reactionary area. And at the same area, cllagene-1 and ALP expression level increased in osteoblast progenitor cells. These data is suggesting that MDSCs emerge to suppress inflammation and support new bone formation. Here, we report, for the first time (to our knowledge), the role of MDSCs in the initiation of bone formation. MDSC appeared at the transition from inflammation to bone making and regulates bone healing by suppressing inflammation.

Exploiting bone niches: progression of disseminated tumor cells to metastasis.

Many solid cancers metastasize to the bone and bone marrow (BM). This process may occur even before the diagnosis of primary tumors, as evidenced by the discovery of disseminated tumor cells (DTCs) in patients without occult malignancies. The cellular fates and metastatic progression of DTCs are determined by complicated interactions between cancer cells and BM niches. Not surprisingly, these niches also play important roles in normal biology, including homeostasis and turnover of skeletal and hematopoiesis systems. In this Review, we summarize recent findings on functions of BM niches in bone metastasis (BoMet), particularly during the early stage of colonization. In light of the rich knowledge of hematopoiesis and osteogenesis, we highlight how DTCs may progress into overt BoMet by taking advantage of niche cells and their activities in tissue turnover, especially those related to immunomodulation and bone repair.

Evidence of macrophage modulation in the mouse pubic symphysis remodeling during the end of first pregnancy and postpartum.

In mouse pregnancy, pubic symphysis (PS) remodels into an elastic interpubic ligament (IpL) in a temporally regulated process to provide safe delivery. It restores at postpartum to assure reproductive tract homeostasis. Recently, macrophage localization in the IpL and dynamic changes in the expression of inflammatory mediators observed from the end of pregnancy (D18, D19) to early days postpartum (1dpp, 3dpp) highlighted the necessity of the identification of the key molecules involved in innate immune processes in PS remodeling. Therefore, this study uses morphological and high-sensitivity molecular techniques to identify both macrophage association with extracellular matrix (ECM) remodeling and the immunological processes involved in PS changes from D18 to 3dpp. Results showed macrophage association with active gelatinases and ECM components and 25 differentially expressed genes (DEGs) related to macrophage activities in interpubic tissues from D18 to 3dpp. Additionally, microarray and proteomic analysis showed a significant association of interpubic tissue DEGs with complement system activation and differentially expressed proteins (DEPs) with phagocytosis, highlighting the involvement of macrophage-related activities in mouse PS remodeling. Therefore, the findings suggest that PS ECM remodeling is associated with evidence of macrophage modulation that ensures both IpL relaxation and fast PS recovery postpartum for first labor.

The role of macrophages in osteoarthritis and cartilage repair.

Osteoarthritis (OA) is a family of degenerative diseases affecting multiple joint tissues. Despite the diverse etiology and pathogenesis of OA, increasing evidence suggests that macrophages can play a significant role in modulating joint inflammation, and thus OA severity, via various secreted mediators. Recent advances in next-generation sequencing technologies coupled with proteomic and epigenetic tools have greatly facilitated research to elucidate the embryonic origin of macrophages in various tissues including joint synovium. Furthermore, scientists have now begun to appreciate that macrophage polarization can span beyond the conventionally recognized binary states (i.e., pro-inflammatory M1-like vs anti-inflammatory M2-like) and may encompass a broad spectrum of phenotypes. Although the presence of these cells has been shown in multiple joint tissues, additional mechanistic studies are required to provide a comprehensive understanding of the precise role of these diverse macrophage populations in OA onset and progression. New approaches that can modulate macrophages into desired functional phenotypes may provide novel therapeutic strategies for preventing OA or enhancing cartilage repair and regeneration.

Immune Function and Diversity of Osteoclasts in Normal and Pathological Conditions.

Osteoclasts (OCLs) are key players in controlling bone remodeling. Modifications in their differentiation or bone resorbing activity are associated with a number of pathologies ranging from osteopetrosis to osteoporosis, chronic inflammation and cancer, that are all characterized by immunological alterations. Therefore, the 2000s were marked by the emergence of osteoimmunology and by a growing number of studies focused on the control of OCL differentiation and function by the immune system. At the same time, it was discovered that OCLs are much more than bone resorbing cells. As monocytic lineage-derived cells, they belong to a family of cells that displays a wide heterogeneity and plasticity and that is involved in phagocytosis and innate immune responses. However, while OCLs have been extensively studied for their bone resorption capacity, their implication as immune cells was neglected for a long time. In recent years, new evidence pointed out that OCLs play important roles in the modulation of immune responses toward immune suppression or inflammation. They unlocked their capacity to modulate T cell activation, to efficiently process and present antigens as well as their ability to activate T cell responses in an antigen-dependent manner. Moreover, similar to other monocytic lineage cells such as macrophages, monocytes and dendritic cells, OCLs display a phenotypic and functional plasticity participating to their anti-inflammatory or pro-inflammatory effect depending on their cell origin and environment. This review will address this novel vision of the OCL, not only as a phagocyte specialized in bone resorption, but also as innate immune cell participating in the control of immune responses.

S1P-S1PR1 Signaling: the "Sphinx" in Osteoimmunology.

The fundamental interaction between the immune and skeletal systems, termed as osteoimmunology, has been demonstrated to play indispensable roles in the maintenance of balance between bone resorption and formation. The pleiotropic sphingolipid metabolite, sphingosine 1-phosphate (S1P), together with its cognate receptor, sphingosine-1-phosphate receptor-1 (S1PR1), are known as key players in osteoimmunology due to the regulation on both immune system and bone remodeling. The role of S1P-S1PR1 signaling in bone remodeling can be directly targeting both osteoclastogenesis and osteogenesis. Meanwhile, inflammatory cell function and polarization in both adaptive immune (T cell subsets) and innate immune cells (macrophages) are also regulated by this signaling axis, suggesting that S1P-S1PR1 signaling could aslo indirectly regulate bone remodeling via modulating the immune system. Therefore, it could be likely that S1P-S1PR1 signaling might take part in the maintenance of continuous bone turnover under physiological conditions, while lead to the pathogenesis of bone deformities during inflammation. In this review, we summarized the immunological regulation of S1P-S1PR1 signal axis during bone remodeling with an emphasis on how osteo-immune regulators are affected by inflammation, an issue with relevance to chronical bone disorders such as rheumatoid arthritis, spondyloarthritis and periodontitis.

Dynamics of Bone Cell Interactions and Differential Responses to PTH and Antibody-Based Therapies.

We propose a mathematical model describing the dynamics of osteoblasts and osteoclasts in bone remodeling. The goal of this work is to develop an integrated modeling framework for bone remodeling and bone cell signaling dynamics that could be used to explore qualitatively combination treatments for osteoporosis in humans. The model has been calibrated using 57 checks from the literature. Specific global optimization methods based on qualitative objectives have been developed to perform the model calibration. We also added pharmacokinetics representations of three drugs to the model, which are teriparatide (PTH(1-34)), denosumab (a RANKL antibody) and romosozumab (a sclerostin antibody), achieving excellent goodness-of-fit of human clinical data. The model reproduces the paradoxical effects of PTH on the bone mass, where continuous administration of PTH results in bone loss but intermittent administration of PTH leads to bone gain, thus proposing an explanation of this phenomenon. We used the model to simulate different categories of osteoporosis. The main attributes of each disease are qualitatively well captured by the model, for example changes in bone turnover in the disease states. We explored dosing regimens for each disease based on the combination of denosumab and romosozumab, identifying adequate ratios and doses of both drugs for subpopulations of patients in function of categories of osteoporosis and the degree of severity of the disease.

Osteoimmunology.

Bone is a crucial element of the skeletal-locomotor system, but also functions as an immunological organ that harbors hematopoietic stem cells (HSCs) and immune progenitor cells. Additionally, the skeletal and immune systems share a number of regulatory molecules, including cytokines and signaling molecules. Osteoimmunology was created as an interdisciplinary field to explore the shared molecules and interactions between the skeletal and immune systems. In particular, the importance of an inseparable link between the two systems has been highlighted by studies on the pathogenesis of rheumatoid arthritis (RA), in which pathogenic helper T cells induce the progressive destruction of multiple joints through aberrant expression of receptor activator of nuclear factor (NF)-κB ligand (RANKL). The conceptual bridge of osteoimmunology provides not only a novel framework for understanding these biological systems but also a molecular basis for the development of therapeutic approaches for diseases of bone and/or the immune system.

Current Understanding of the Pathophysiology of Osteonecrosis of the Jaw.

PURPOSE OF REVIEW: Osteonecrosis of the jaw (ONJ) is a rare and severe necrotic bone disease reflecting a compromise in the body's osseous healing mechanisms and unique to the craniofacial region. Antiresorptive and antiangiogenic medications have been suggested to be associated with the occurrence of ONJ; yet, the pathophysiology of this disease has not been fully elucidated. This article raises the current theories underlying the pathophysiology of ONJ. RECENT FINDINGS: The proposed mechanisms highlight the unique localization of ONJ. The evidence-based mechanisms of ONJ pathogenesis include disturbed bone remodeling, inflammation or infection, altered immunity, soft tissue toxicity, and angiogenesis inhibition. The role of dental infections and the oral microbiome is central to ONJ, and systemic conditions such as rheumatoid arthritis and diabetes mellitus contribute through their impact on immune resiliency. Current experimental studies on mechanisms of ONJ are summarized. The definitive pathophysiology is as yet unclear. Recent studies are beginning to clarify the relative importance of the proposed mechanisms. A better understanding of osteoimmunology and the relationship of angiogenesis to the development of ONJ is needed along with detailed studies of the impact of drug holidays on the clinical condition of ONJ.

The positive effects of secreting cytokines IL-17 and IFN-γ on the early-stage differentiation and negative effects on the calcification of primary osteoblasts in vitro.

OBJECTIVE: Interleukin-17 (IL-17) and interferon-gamma (IFN-γ) are all pro-inflammatory cytokines produced by specific subsets of T-cells and are also considered crucial regulators in bone remodeling, but their effects on osteogenesis have not been carefully studied. So, this study aimed to investigate the effects of secreting cytokines IL-17 and IFN-γ on the osteogenesis of primary osteoblasts and to clarify the potential roles of the related Janus activated kinase 2 (JAK2) and downstream signal transducer and activator of transcription 3 (STAT3) signaling pathway in bone remodeling. METHODS: The proliferation of osteoblasts was evaluated by MTT assay. Osteogenic activity was tested by alkaline phosphatase (ALP) activity assay and alizarin red staining. The mRNA levels of ALP, osteocalcin, osteoprotegerin (OPG), Runt-related transcription factor 2 (Runx2) and receptor activator of nuclear factor-kappa B ligand (RANKL) were also measured by real-time quantitative PCR. The JAK2-STAT3 pathway was evaluated by Western blot. RESULTS: Osteoblasts showed no obvious proliferation when treated with IL-17 and/or IFN-γ, but higher ALP activities were observed in primary osteoblasts treated with IL-17 or IL-17 + IFN-γ in induction medium. We also found that IL-17 could promote the gene expression of Alp, Runx2, Osteocalcin, Opg, and Rankl, while IFN-γ might attenuate this effect. Nevertheless, IL-17 and IFN-γ exhibited an inhibitory effect on the calcification of primary osteoblasts. We also found that IL-17 could directly facilitate RANKL expressions by JAK2-STAT3 pathway. CONCLUSION: The positive effects of IL-17 and IFN-γ on the early-stage differentiation and the negative effects on the calcification of murine calvarial osteoblasts contribute to our understanding of the role and interaction of inflammatory factors in the bone remodeling and as fundamental mechanisms involved in the destruction of alveolar bone.

Altered Bone Remodeling in Psoriatic Disease: New Insights and Future Directions.

Psoriatic arthritis (PsA) is an inflammatory rheumatic disorder that occurs in patients with psoriasis and predominantly affects musculoskeletal structures, skin, and nails. The etiology of PsA is not well understood but evidence supports an interplay of genetic, immunologic, and environmental factors which promote pathological bone remodeling and joint damage in PsA. Localized and systemic bone loss due to increased activity of osteoclasts is well established in PsA based on animal models and translational studies. In contrast, the mechanisms responsible for pathological bone remodeling in PsA remain enigmatic although new candidate molecules and pathways have been identified. Recent reports have revealed novel findings related to bone erosion and pathologic bone formation in PsA. Many associated risk factors and contributing molecular mechanisms have also been identified. In this review, we discuss new developments in the field, point out unresolved questions regarding the pathogenetic origins of the wide array of bone phenotypes in PsA, and discuss new directions for investigation.

Impairment of Bone Remodeling in LIGHT/TNFSF14-Deficient Mice.

Multiple cytokines produced by immune cells induce remodeling and aid in maintaining bone homeostasis through differentiation of bone-forming osteoblasts and bone-resorbing osteoclasts. Here, we investigate bone remodeling controlled by the tumor necrosis factor (TNF) superfamily cytokine LIGHT. LIGHT-deficient mice (Tnfsf14-/- ) exhibit spine deformity and reduced femoral cancellous bone mass associated with an increase in the osteoclast number and a slight decrease of osteoblasts compared with WT mice. The effect of LIGHT in bone cells can be direct or indirect, mediated by both the low expression of the anti-osteoclastogenic osteoprotegerin (OPG) in B and T cells and reduced levels of the pro-osteoblastogenic Wnt10b in CD8+ T cells in Tnfsf14-/- mice. LIGHT stimulation increases OPG levels in B, CD8+ T, and osteoblastic cells, as well as Wnt10b expression in CD8+ T cells. The high bone mass in Light and T- and B-cell-deficient mice (Rag- /Tnfsf14- ) supports the cooperative role of the immune system in bone homeostasis. These results implicate LIGHT as a potential target in bone disease. © 2017 American Society for Bone and Mineral Research.

Distinct Effects of IL-6 Classic and Trans-Signaling in Bone Fracture Healing.

Bone healing is a complex process with closely linked phases of inflammation, regeneration, and remodeling. IL-6 may crucially regulate this process; however, the underlying mechanisms are unclear. IL-6 signals are transmitted via the transmembrane glycoprotein 130 by two distinct mechanisms: classic signaling using the membrane-anchored IL-6 receptor and trans-signaling using its soluble form. Herein, we investigated the hypothesis that IL-6 classic and trans-signaling have different functions during bone healing. To investigate fracture healing, 12-week-old C57BL/6J mice underwent a femur osteotomy. To study the function of IL-6 during the inflammatory phase, either an anti-IL-6 antibody, which inhibits IL-6 classic and trans-signaling, or soluble glycoprotein 130 fusion protein, which selectively blocks trans-signaling, was injected after 30 minutes and 48 hours. To analyze IL-6 effects in the repair phase, compounds were injected from day 7 onwards. Global IL-6 inhibition in the early phase after fracture reduced systemic inflammation, the recruitment of immune cells, and bone regeneration, resulting in delayed fracture healing. Global IL-6 inhibition during the repair phase disturbed bone formation and remodeling. In contrast, inhibition of IL-6 trans-signaling exerted minor effects on the immune response and did not influence bone repair, suggesting that the classic pathway accounts for most of the effects observed after global IL-6 inhibition. Our results reveal that IL-6 classic signaling, but not IL-6 trans-signaling, is essential for bone repair.

Bone and the Immune System.

Osteoporosis increases fracture risk, a cause of crippling morbidity and mortality. The immunoskeletal interface (ISI) is a centralization of cell and cytokine effectors shared between skeletal and immune systems. Consequently, the immune system mediates powerful effects on bone turnover. Physiologically, B cells secrete osteoprotegerin (OPG), a potent anti-osteoclastogenic factor that preserves bone mass. However, activated T cells and B cells secrete pro-osteoclastogenic factors including receptor activator of Nuclear factor-kappaB (NF-kB) ligand (RANKL), Interleukin (IL)-17A, and tumor necrosis factor (TNF)-α promoting bone loss in inflammatory states such as rheumatoid arthritis. Recently, ISI disruption has been linked to osteoporosis in human immunodeficiency virus (HIV) infection/acquired immunodeficiency syndrome (AIDS), where elevated B cell RANKL and diminished OPG drive bone resorption. HIV-antiretroviral therapy paradoxically intensifies bone loss during disease reversal, as immune reconstitution produces osteoclastogenic cytokines. Interestingly, in estrogen deficiency, activated T cells secrete RANKL, TNF, and IL-17A that amplify bone resorption and contribute to postmenopausal osteoporosis. T cell-produced TNF and IL-17A further contribute to bone loss in hyperparathyroidism, while T cell production of the anabolic Wingless integration site (Wnt) ligand, Wnt10b, promotes bone formation in response to anabolic parathyroid hormone and the immunomodulatory costimulation inhibitor cytotoxic T lymphocyte-associated protein-4-IgG (abatacept). These findings provide a window into the workings of the ISI and suggest novel targets for future therapeutic interventions to reduce fracture risk.

Osteoimmunology in Bone Fracture Healing.

PURPOSE OF REVIEW: In the process of bone fracture healing, inflammation is thought to be an essential process that precedes bone formation and remodeling. We review recent studies on bone fracture healing from an osteoimmunological point of view. RECENT FINDINGS: Based on previous observations that many types of immune cells infiltrate into the bone injury site and release a variety of molecules, recent studies have addressed the roles of specific immune cell subsets. Macrophages and interleukin (IL)-17-producing γδ T cells enhance bone healing, whereas CD8+ T cells impair bone repair. Additionally, IL-10-producing B cells may contribute to bone healing by suppressing excessive and/or prolonged inflammation. Although the involvement of other cells and molecules has been suggested, the precise underlying mechanisms remain elusive. Accumulating evidence has begun to reveal the deeper picture of bone fracture healing. Further studies are required for the development of novel therapeutic strategies for bone fracture.