ABSTRACT Osteocytes appear to mobilize calcium within minutes in response to PTH injections; we have previously shown that osteocytes remove their perilacunar matrix during lactation through activation of the PTH type 1 receptor. Mechanisms utilized by osteocytes to mobilize calcium are unknown but we hypothesized that the molecular components may be similar to those used by osteoclasts. Here we show, using IDG-SW3 cells that ATP6V0D2, an essential component of vacuolar ATPase in osteoclasts, and other genes associated with osteoclastic bone resorption, increase with osteoblast to osteocyte differentiation. Furthermore, PTHrP increases ATP6V0D2 expression and induces proton generation by primary osteocytes, which is blocked by bafilomycin, a vacuolar ATPase inhibitor. These in vitro proton measurements raised the question of osteocyte viability in an acidic environment. Interestingly, osteocytes, showed enhanced viability at pH as low as 5 compared to osteoblasts and fibroblasts in vitro. To study in vivo acidification by osteocytes, virgin and lactating CD1 mice on a low calcium diet were injected with the pH indicator dye, acridine orange, and their osteocyte lacuno-canalicular system imaged by confocal microscopy. Lower pH was observed in lactating compared to virgin animals. In addition, a novel transgenic mouse line with a topaz variant of green fluorescent protein (GFPtpz)-tagged collagen α2(I) chain was used. Instead of the expected reduction in GFP-fluorescence only in the perilacunar matrix, reduced fluorescence was observed in the entire bone matrix of lactating mice. Based on our experiments showing quenching of GFP in vitro, we propose that the observed reduction in GFP fluorescence in lactating mice is due to quenching of GFP by the acidic pH generated by osteocytes. Together these findings provide novel mechanistic insight into how osteocytes remove calcium from their perilacunar/pericanalicular matrices through active acidification of their microenvironment and show that osteocytes, like osteoclasts, are resistant to the negative effects of acid on viability. © 2017 American Society for Bone and Mineral Research.
The amazing osteocyte
Abstract The last decade has provided a virtual explosion of data on the molecular biology and function of osteocytes. Far from being the “passive placeholder in bone,” this cell has been found to have numerous functions, such as acting as an orchestrator of bone remodeling through regulation of both osteoclast and osteoblast activity and also functioning as an endocrine cell. The osteocyte is a source of soluble factors not only to target cells on the bone surface but also to target distant organs, such as kidney, muscle, and other tissues. This cell plays a role in both phosphate metabolism and calcium availability and can remodel its perilacunar matrix. Osteocytes compose 90% to 95% of all bone cells in adult bone and are the longest lived bone cell, up to decades within their mineralized environment. As we age, these cells die, leaving behind empty lacunae that frequently micropetrose. In aged bone such as osteonecrotic bone, empty lacunae are associated with reduced remodeling. Inflammatory factors such as tumor necrosis factor and glucocorticoids used to treat inflammatory disease induce osteocyte cell death, but by different mechanisms with potentially different outcomes. Therefore, healthy, viable osteocytes are necessary for proper functionality of bone and other organs. © 2011 American Society for Bone and Mineral Research.
TRANCE Is Necessary and Sufficient for Osteoblast-mediated Activation of Bone Resorption in Osteoclasts
TRANCE (tumor necrosis factor–related activation-induced cytokine) is a recently described member of the tumor necrosis factor superfamily that stimulates dendritic cell survival and has also been found to induce osteoclastic differentiation from hemopoietic precursors. However, its effects on mature osteoclasts have not been defined. It has long been recognized that stimulation of osteoclasts by agents such as parathyroid hormone (PTH) occurs through a hormonal interaction with osteoblastic cells, which are thereby induced to activate osteoclasts. To determine whether TRANCE accounts for this activity, we tested its effects on mature osteoclasts. TRANCE rapidly induced a dramatic change in osteoclast motility and spreading and inhibited apoptosis. In populations of osteoclasts that were unresponsive to PTH, TRANCE caused activation of bone resorption equivalent to that induced by PTH in the presence of osteoblastic cells. Moreover, osteoblast-mediated stimulation of bone resorption was abrogated by soluble TRANCE receptor and by the soluble decoy receptor osteoprotegerin (OPG), and stimulation of isolated osteoclasts by TRANCE was neutralized by OPG. Thus, TRANCE expression by osteoblasts appears to be both necessary and sufficient for hormone-mediated activation of mature osteoclasts, and TRANCE-R is likely to be a receptor for signal transduction for activation of the osteoclast and its survival.
The osteocyte as a signaling cell
Osteocytes, former osteoblasts encapsulated by mineralized bone matrix, are far from being passive and metabolically inactive bone cells. Instead, osteocytes are multifunctional and dynamic cells capable of integrating hormonal and mechanical signals and transmitting them to effector cells in bone and in distant tissues. Osteocytes are a major source of molecules that regulate bone homeostasis by integrating both mechanical cues and hormonal signals that coordinate the differentiation and function of osteoclasts and osteoblasts. Osteocyte function is altered in both rare and common bone diseases, suggesting that osteocyte dysfunction is directly involved in the pathophysiology of several disorders affecting the skeleton. Advances in osteocyte biology initiated the development of novel therapeutics interfering with osteocyte-secreted molecules. Moreover, osteocytes are targets and key distributors of biological signals mediating the beneficial effects of several bone therapeutics used in the clinic. Here we review the most recent discoveries in osteocyte biology demonstrating that osteocytes regulate bone homeostasis and bone marrow fat via paracrine signaling, influence body composition and energy metabolism via endocrine signaling, and contribute to the damaging effects of diabetes mellitus and hematologic and metastatic cancers in the skeleton.
Control of Bone Matrix Properties by Osteocytes
Osteocytes make up 90-95% of the cellular content of bone and form a rich dendritic network with a vastly greater surface area than either osteoblasts or osteoclasts. Osteocytes are well positioned to play a role in bone homeostasis by interacting directly with the matrix; however, the ability for these cells to modify bone matrix remains incompletely understood. With techniques for examining the nano- and microstructure of bone matrix components including hydroxyapatite and type I collagen becoming more widespread, there is great potential to uncover novel roles for the osteocyte in maintaining bone quality. In this review, we begin with an overview of osteocyte biology and the lacunar-canalicular system. Next, we describe recent findings from in vitro models of osteocytes, focusing on the transitions in cellular phenotype as they mature. Finally, we describe historical and current research on matrix alteration by osteocytes in vivo, focusing on the exciting potential for osteocytes to directly form, degrade, and modify the mineral and collagen in their surrounding matrix.
Regulation of Bone Remodeling by Parathyroid Hormone
Parathyroid hormone (PTH) exerts profound effects on skeletal homeostasis through multiple cellular and molecular mechanisms. Continuous hyperparathyroidism causes net loss of bone mass, despite accelerating bone formation by osteoblasts. Intermittent treatment with PTH analogs represents the only Food and Drug Administration (FDA)-approved bone anabolic osteoporosis treatment strategy. Functional PTH receptors are present on cells of the osteoblast lineage, ranging from early skeletal stem cells to matrix-embedded osteocytes. In addition, bone remodeling by osteoclasts liberates latent growth factors present within bone matrix. Here, we will provide an overview of the multiple cellular and molecular mechanisms through which PTH influences bone homeostasis. Notably, net skeletal effects of continuous versus intermittent can differ significantly. Where possible, we will highlight mechanisms through which continuous hyperparathyroidism leads to bone loss, and through which intermittent hyperparathyroidism boosts bone mass. Given the therapeutic usage of intermittent PTH (iPTH) treatment for osteoporosis, particular attention will be paid toward mechanisms underlying the bone anabolic effects of once daily PTH administration.
Mathematical Model of Paracrine Interactions between Osteoclasts and Osteoblasts Predicts Anabolic Action of Parathyroid Hormone on Bone
Abstract. To restore falling plasma calcium levels, PTH promotes calcium liberation from bone. PTH targets bone-forming cells, osteoblasts, to increase exp
Osteocyte-Related Cytokines Regulate Osteoclast Formation and Bone Resorption
The process of bone remodeling is the result of the regulated balance between bone cell populations, namely bone-forming osteoblasts, bone-resorbing osteoclasts, and the osteocyte, the mechanosensory cell type. Osteoclasts derived from the hematopoietic stem cell lineage are the principal cells involved in bone resorption. In osteolytic diseases such as rheumatoid arthritis, periodontitis, and osteoporosis, the balance is lost and changes in favor of bone resorption. Therefore, it is vital to elucidate the mechanisms of osteoclast formation and bone resorption. It has been reported that osteocytes express Receptor activator of nuclear factor κΒ ligand (RANKL), an essential factor for osteoclast formation. RANKL secreted by osteocytes is the most important factor for physiologically supported osteoclast formation in the developing skeleton and in pathological bone resorption such as experimental periodontal bone loss. TNF-α directly enhances RANKL expression in osteocytes and promotes osteoclast formation. Moreover, TNF-α enhances sclerostin expression in osteocytes, which also increases osteoclast formation. These findings suggest that osteocyte-related cytokines act directly to enhance osteoclast formation and bone resorption. In this review, we outline the most recent knowledge concerning bone resorption-related cytokines and discuss the osteocyte as the master regulator of bone resorption and effector in osteoclast formation.
The Development of Molecular Biology of Osteoporosis
Osteoporosis is one of the major bone disorders that affects both women and men, and causes bone deterioration and bone strength. Bone remodeling maintains bone mass and mineral homeostasis through the balanced action of osteoblasts and osteoclasts, which are responsible for bone formation and bone resorption, respectively. The imbalance in bone remodeling is known to be the main cause of osteoporosis. The imbalance can be the result of the action of various molecules produced by one bone cell that acts on other bone cells and influence cell activity. The understanding of the effect of these molecules on bone can help identify new targets and therapeutics to prevent and treat bone disorders. In this article, we have focused on molecules that are produced by osteoblasts, osteocytes, and osteoclasts and their mechanism of action on these cells. We have also summarized the different pharmacological osteoporosis treatments that target different molecular aspects of these bone cells to minimize osteoporosis.
Biology of Bone Tissue: Structure, Function, and Factors That Influence Bone Cells
Bone tissue is continuously remodeled through the concerted actions of bone cells, which include bone resorption by osteoclasts and bone formation by osteoblasts, whereas osteocytes act as mechanosensors and orchestrators of the bone remodeling process. This process is under the control of local (e.g., growth factors and cytokines) and systemic (e.g., calcitonin and estrogens) factors that all together contribute for bone homeostasis. An imbalance between bone resorption and formation can result in bone diseases including osteoporosis. Recently, it has been recognized that, during bone remodeling, there are an intricate communication among bone cells. For instance, the coupling from bone resorption to bone formation is achieved by interaction between osteoclasts and osteoblasts. Moreover, osteocytes produce factors that influence osteoblast and osteoclast activities, whereas osteocyte apoptosis is followed by osteoclastic bone resorption. The increasing knowledge about the structure and functions of bone cells contributed to a better understanding of bone biology. It has been suggested that there is a complex communication between bone cells and other organs, indicating the dynamic nature of bone tissue. In this review, we discuss the current data about the structure and functions of bone cells and the factors that influence bone remodeling.
The Effect of Inflammation on Bone
Bone remodeling is the continual process to renew the adult skeleton through the sequential action of osteoblasts and osteoclasts. Nuclear factor RANK, an osteoclast receptor, and its ligand RANKL, expressed on the surface of osteoblasts, result in coordinated control of bone remodeling. Inflammation, a feature of illness and injury, plays a distinct role in skewing this process toward resorption. It does so via the interaction of inflammatory mediators and their related peptides with osteoblasts and osteoclasts, to alter the expression of RANK and RANKL. Such chemical mediators include TNFa, glucocorticoids, histamine, bradykinin, PGE2, systemic RANKL from immune cells, and interleukins 1 and 6. A thorough understanding of bone response to injury and disease, and ability to detect such biomarkers, as well as imaging to identify early structural and mechanical property changes in bone architecture, is important in improving management and outcomes of bone related pathology. While gut health and vitamin and mineral availability appear vitally important, there are very few intervention strategies currently available to favorably alter bone pathology induced by inflammation. Further research is required in this field to advance early detection and treatments.
Loss of Bone and Wnt10b Expression in Male Type 1 Diabetic Mice Is Blocked by the Probiotic Lactobacillus reuteri
Type 1 diabetes (T1D)–induced osteoporosis is characterized by a predominant suppression of osteoblast number and activity, as well as increased bone marrow adiposity but no change in osteoclast activity. The fundamental mechanisms and alternative anabolic treatments (with few side effects) for T1D bone loss remain undetermined. Recent studies by our laboratory and others indicate that probiotics can benefit bone health. Here, we demonstrate that Lactobacillus reuteri, a probiotic with anti-inflammatory and bone health properties, prevents T1D-induced bone loss and marrow adiposity in mice. We further found that L. reuteri treatment prevented the suppression of Wnt10b in T1D bone. Consistent with a role for attenuated bone Wnt10b expression in T1D osteoporosis, we observed that bone-specific Wnt10b transgenic mice are protected from T1D bone loss. To examine the mechanisms of this protection, we focused on TNF-α, a cytokine up-regulated in T1D that causes suppression of osteoblast Wnt10b expression in vitro. Addition of L. reuteri prevented TNF-α–mediated suppression of Wnt10b and osteoblast maturation markers. Taken together, our findings reveal a mechanism by which T1D causes bone loss and open new avenues for use of probiotics to benefit the bone.
Molecular Signaling Pathways and Essential Metabolic Elements in Bone Remodeling: An Implication of Therapeutic Targets for Bone Diseases
Bone is one of the dynamic tissues in the human body that undergoes continuous remodelling through subsequent actions of bone cells, osteoclasts, and osteoblasts. Several signal transduction pathways are involved in the transition of mesenchymal stem cells into osteoblasts. These primarily include Runx2, ATF4, Wnt signaling and sympathetic signalling. The differentiation of osteoclasts is controlled by M-CSF, RANKL, and costimulatory signalling. It is well known that bone remodelling is regulated through receptor activator of nuclear factor-kappa B ligand followed by the binding to RANK, which eventually induces the differentiation of osteoclasts. The resorbing osteoclasts secrete TRAP, cathepsin K, MMP-9 and gelatinase to digest the proteinaceous matrix of type I collagen and form a saucer-shaped lacuna along with resorption tunnels in the trabecular bone. Osteoblasts secrete a soluble decoy receptor, osteoprotegerin that prevents the binding of RANK/RANKL and thus moderating osteoclastogenesis. Moreover, bone homeostasis is also regulated by several growth factors, cytokines, calciotropic hormones, parathyroid hormone and sex steroids. The current review presents a correlation of the probable molecular targets underlying the regulation of bone mass and the role of essential metabolic elements in bone remodelling. Targeting these signaling pathways may help design newer therapies for treating bone diseases.
Dynamics of bone cell signaling and PTH treatments of osteoporosis
In this paper we analyze and generalize the dynamical system introduced by Lemaire and co-workers [14] as a model of cell signaling in bone remodeling. We show that for large classes of parameter values, including the physically-realistic baseline values, the system has a unique physically relevant equilibrium which is a global attractor. We generalize that model, minimally, to incorporate a mechanism by which parathyroid hormone (PTH) retards osteoblast apoptosis. We show that with this mechanism, which is a simplified version of that proposed by Bellido and co-workers [4], the model exhibits a well-known phenomenon that has puzzled researchers: the system responds catabolically to the continuous administration of PTH andally to appropriately pulsed administration of PTH.
Osteoblast-Osteoclast Communication and Bone Homeostasis
Bone remodeling is tightly regulated by a cross-talk between bone-forming osteoblasts and bone-resorbing osteoclasts. Osteoblasts and osteoclasts communicate with each other to regulate cellular behavior, survival and differentiation through direct cell-to-cell contact or through secretory proteins. A direct interaction between osteoblasts and osteoclasts allows bidirectional transduction of activation signals through EFNB2-EPHB4, FASL-FAS or SEMA3A-NRP1, regulating differentiation and survival of osteoblasts or osteoclasts. Alternatively, osteoblasts produce a range of different secretory molecules, including M-CSF, RANKL/OPG, WNT5A, and WNT16, that promote or suppress osteoclast differentiation and development. Osteoclasts also influence osteoblast formation and differentiation through secretion of soluble factors, including S1P, SEMA4D, CTHRC1 and C3. Here we review the current knowledge regarding membrane bound- and soluble factors governing cross-talk between osteoblasts and osteoclasts.
The bone remodelling cycle
The bone remodelling cycle replaces old and damaged bone and is a highly regulated, lifelong process essential for preserving bone integrity and maintaining mineral homeostasis. During the bone remodelling cycle, osteoclastic resorption is tightly coupled to osteoblastic bone formation. The remodelling cycle occurs within the basic multicellular unit and comprises five co-ordinated steps; activation, resorption, reversal, formation and termination. These steps occur simultaneously but asynchronously at multiple different locations within the skeleton. Study of rare human bone disease and animal models have helped to elucidate the cellular and molecular mechanisms that regulate the bone remodelling cycle. The key signalling pathways controlling osteoclastic bone resorption and osteoblastic bone formation are receptor activator of nuclear factor-κB (RANK)/RANK ligand/osteoprotegerin and canonical Wnt signalling. Cytokines, growth factors and prostaglandins act as paracrine regulators of the cycle, whereas endocrine regulators include parathyroid hormone, vitamin D, calcitonin, growth hormone, glucocorticoids, sex hormones, and thyroid hormone. Disruption of the bone remodelling cycle and any resulting imbalance between bone resorption and formation leads to metabolic bone disease, most commonly osteoporosis. The advances in understanding the cellular and molecular mechanisms underlying bone remodelling have also provided targets for pharmacological interventions which include antiresorptive and anabolic therapies. This review will describe the remodelling process and its regulation, discuss osteoporosis and summarize the commonest pharmacological interventions used in its management.
Lactulose Suppresses Osteoclastogenesis and Ameliorates Estrogen Deficiency-Induced Bone Loss in Mice
<p>Postmenopausal osteoporosis is characterized by excess osteoclastogenesis which leads to net bone loss and brittle fractures. Studies have demonstrated that estrogen deficiency-associated bone loss is microbiota-dependent and could be prevented by probiotics and prebiotics. In this study, we report that orally administered lactulose (20 g/kg, 6 weeks) orally administered significantly inhibited osteoclastogenesis, bone resorption, and prevented ovariectomy (OVX)-induced bone loss in mice. Lactulose increased intestinal <i>Claudin 2, 3 and 15,</i> compared to the OVX group, and lowered pro-osteoclastogenic cytokines levels including tumor necrosis factor-α, interleukin(IL)-6, receptor activator of nuclear factor kappa-Β ligand (RANKL), and IL-17 as well as increased the anti-inflammatory cytokine IL-10 in the intestine, peripheral blood, and bone marrow. Lactulose significantly preserved the number of Foxp3<sup>+</sup> Treg cells in the intestines compared with that in OVX mice. Lactulose altered the composition of intestinal microbiota measured by 16s rDNA sequencing and increased intestinal and serum short-chain fatty acids (SCFAs) levels including acetate, propionate and butyrate which were decreased in OVX mice as measured by gas chromatography. Oral administration of lactulose for 2 weeks significantly lowered the level of bone resorption marker C-telopeptide of type 1 collagen-1 in healthy male young volunteers (aging 20-25 years). In conclusion, lactulose inhibited osteoclastogenesis and bone resorption by altering the intestinal microbiota and increasing SCFAs. Lactulose could serve as an ideal therapeutic agent for postmenopausal osteoporosis.</p>
