transforming-growth-factor-beta and Osteolysis

Calcinosis cutis, defined as sub-epidermal deposition of calcium salts, is a major clinical problem in patients with SSc, affecting 20-40% of patients. A number of recognized factors associated with calcinosis have been identified, including disease duration, digital ischaemia and acro-osteolysis. Yet, to date, the pathogenesis of SSc-related calcinosis remains unknown, and currently there is no effective disease-modifying pharmacotherapy. Following onset of SSc, there are marked changes in the extracellular matrix (ECM) of the skin, notably a breakdown in the microfibrillar network and accumulation of type I collagen. Our hypothesis is that these pathological changes reflect a changing cellular phenotype and result in a primed microenvironment for soft tissue calcification, with SSc fibroblasts adopting a pro-osteogenic profile, and specific driving forces promoting tissue mineralization. Considering the role of the ECM in disease progression may help elucidate the mechanism(s) behind SSc-related calcinosis and inform the development of future therapeutic interventions.

Topics: Calcinosis; Cell Differentiation; Cell Hypoxia; Cellular Microenvironment; Collagen Type I; Disease Progression; Elastin; Extracellular Matrix; Fibrillin-1; Fibroblasts; Glucose Transporter Type 1; Humans; Mesenchymal Stem Cells; Myofibroblasts; Osteoblasts; Osteogenesis; Osteolysis; Phenotype; Phosphates; Scleroderma, Systemic; Skin; Transforming Growth Factor beta; Vascular Endothelial Growth Factor A

Bone remodeling is a delicate balancing act between the bone matrix synthesizing osteoblasts and bone resorbing osteoclasts. Active bone metastases typically subvert this process to generate lesions that are comprised of extensive areas of pathological osteogenesis and osteolysis. The resultant increase in bone matrix remodeling enhances cytokine/growth factor bioavailability thus creating a vicious cycle that stimulates tumor progression. Given the extent of matrix remodeling occurring in the tumor-bone microenvironment, the expression of matrix metalloproteinases (MMPs) would be expected, since collectively they have the ability to degrade all components of the extracellular matrix (ECM). However, in addition to being "matrix bulldozers", MMPs control the bioavailability and bioactivity of factors such as RANKL and TGFβ that have been described as crucial for tumor-bone interaction, thus implicating MMPs as key regulators of the vicious cycle of bone metastases.

Topics: Bone and Bones; Bone Neoplasms; Bone Remodeling; Bone Resorption; Extracellular Matrix; Humans; Matrix Metalloproteinases; Osteoblasts; Osteoclasts; Osteogenesis; Osteolysis; RANK Ligand; Transforming Growth Factor beta; Tumor Microenvironment

In normal bone there are two essential processes of bone turnover, resorption and formation, which are disrupted by bone metastases. Two types of bone metastases are known, i.e. osteolytic lesions with dominant bone resorption and osteosclerotic tumors with enhanced osteoblastic bone formation. Numerous cytokines and growth factors regulate the activity of osteoclasts and/or osteoblasts in endo- or paracrine ways, playing crucial roles in the processes of bone turnover. Bone metastases are often the consequences of certain malignant tumors, such as breast, prostate, lung, and renal cancer. The diagnosis of bone metastasis is essential for a determination of the clinical stage of cancer and appropriate treatment. Tumor markers are useful in diagnosis, prognosis, staging, and, especially, monitoring treatment. Tumor markers are also useful in detecting bone metastases. There is growing evidence that various cytokines, especially M-CSF, TGFbeta, TNFalpha, and IL-6 and IL-7, may be new tumor markers useful in the diagnosis of neoplastic disease. The processes of bone turnover in normal bone and metastatic tumors as well as the significance of the most important cytokines in the development of osteolytic metastases and the possibility of their use in the diagnosis of the most frequent cancers presenting bone metastases are described in this article.

Topics: Biomarkers, Tumor; Bone Neoplasms; Cytokines; Humans; Interleukin-6; Macrophage Colony-Stimulating Factor; Osteolysis; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Breast cancer most frequently causes bone metastases in solid tumors. It has been known that there is a vicious cycle consisting of tumor cells, osteoblasts, osteoclasts and various humoral factors in osteolytic lesions. Although systemic therapy is a main treatment of bone metastases, local therapies, such as radiotherapy and surgical therapy, are also promptly needed when bone-related complications occur. In recent years, anti-osteoclast agents, bisphosphonates significantly contribute to the delay of occurrence of bone-related complications. Postoperative adjuvant therapy significantly reduces the incidence of recurrence in breast cancer patients. Chemotherapy and LH-RH agonists cause ovarian function suppression in premenopausal patients, and aromatase inhibitors cause estrogen deprivation in postmenopausal patients. These effects cause unbalance of bone metabolism, loss of bone density and increase in the incidence of fractures. Improvement of these bone-related adverse effects and careful follow-ups are needed for breast cancer patients.

Topics: Antineoplastic Agents; Antineoplastic Agents, Hormonal; Bone Density Conservation Agents; Bone Neoplasms; Breast Neoplasms; Chemotherapy, Adjuvant; Diphosphonates; Female; Humans; Medroxyprogesterone Acetate; Osteolysis; Parathyroid Hormone-Related Protein; RANK Ligand; Transforming Growth Factor beta

Extensive bone marrow necrosis and symptomatic hypercalcemia have been described independently as rare complications of chronic myeloid leukemia. Here we report a 66-year-old man who developed B cell blastic transformation 10 years after diagnosis of CML in the chronic phase. Extensive bone marrow necrosis and symptomatic hypercalcemia concurrently developed after transformation, with development of disseminated intravascular coagulation and multifocal osteolysis. Most necrotic cells were readily identifiable as blasts. Mediators related to hypercalcemia, including prostaglandin E2, transforming growth factor-alpha and transforming growth factor-beta, were significantly elevated in the serum. As far as we know, this is the first case report of chronic myeloid leukemia concurrently developing bone marrow necrosis and hypercalcemia; this association was not reported in other types of leukemia or bone marrow malignancies.

Topics: Blast Crisis; Bone Marrow; Dinoprostone; Disseminated Intravascular Coagulation; Fatal Outcome; Humans; Hypercalcemia; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Male; Middle Aged; Necrosis; Osteolysis; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Several common cancers often metastasize to the skeleton in advanced disease. Bone metastases are incurable and cause protracted, severe symptoms. Growth of tumor in bone is driven by a vicious cycle: tumor-secreted factors stimulate bone cells, which in turn release growth factors and cytokines. The bone-derived factors fuel the vicious cycle by acting back on the tumor cells. The vicious cycle offers novel targets for the treatment of advanced cancers. Treatments can inhibit bone cells (osteoclasts and osteoblasts) that are stimulated by tumor-secreted factors. Drugs can also inhibit tumor responses to factors enriched in the bone microenvironment, such as transforming growth factor-beta. Animal models show that these approaches, especially combination treatments, can reduce tumor burden. The results suggest a novel paradigm in which tumor growth can be effectively inhibited by drugs that target cells in the bone microenvironment and not the tumor cells themselves.

Topics: Bone Neoplasms; Forecasting; Humans; Models, Biological; Neoplasm Metastasis; Osteoblasts; Osteoclasts; Osteolysis; Transforming Growth Factor beta; Tumor Burden

Metastasis is a final stage of tumor progression. Breast and prostate cancer cells preferentially metastasize to bone, wherein they cause incurable osteolytic and osteoblastic lesions. The bone matrix is rich in factors, such as transforming growth factor-beta and insulin-like growth factors, which are released into the tumor microenvironment by osteolysis. These factors stimulate the growth of tumor cells and alter their phenotype, thus promoting a vicious cycle of metastasis and bone pathology. Physical factors within the bone microenvironment, including low oxygen levels, acidic pH, and high extracellular calcium concentrations, may also enhance tumor growth. These elements of the microenvironment are potential targets for chemotherapeutic intervention to halt tumor growth and suppress bone metastasis.

Topics: Bone Neoplasms; Breast Neoplasms; Female; Humans; Hydrogen-Ion Concentration; Male; Molecular Biology; Osteolysis; Prostatic Neoplasms; Somatomedins; Transforming Growth Factor beta

The most common skeletal complication of breast cancer is osteolytic bone metastasis. Bone metastases are present in 80% of patients with advanced disease and cause significant morbidity. They are most often osteolytic, but can be osteoblastic or mixed. Tumor cells, osteoblasts, osteoclasts and bone matrix are the four components of a vicious cycle necessary for the initiation and development of bone metastases. Tumor cell gene expression is modified by interaction with bone-derived factors. For example, parathyroid hormone related protein (PTHrP), a tumor cell factor, is upregulated by bone-derived transforming growth factor beta (TGFbeta). Tumor cell factors, in turn, act upon bone cells to cause dysregulated bone destruction and formation. PTHrP increases osteoblast expression of RANK (receptor activator of NFkappaB) ligand which, in turn, activates osteoclasts. PTHrP-independent osteolytic factors, such as interleukin [IL]-11 and IL-8, also contribute to the vicious cycle. Other tumor-bone interactions, such as stimulation of tumor-homing through the CXCR4 chemokine receptor by its bone-derived ligand stromal-derived factor-1 (SDF-1), may be responsible for the site-specific predilection of breast cancer for bone. These factors and their roles in fueling the vicious cycle may identify novel targets for therapies to prevent metastasis.

Topics: Animals; Bone Neoplasms; Bone Resorption; Breast Neoplasms; Female; Humans; Osteolysis; Parathyroid Hormone-Related Protein; Signal Transduction; Trans-Activators; Transforming Growth Factor beta

Breast cancers frequently metastasize to the skeleton and cause bone destruction. Tumor cells secrete factors that stimulate osteoclasts. The consequent osteolytic resorption releases active factors from the bone matrix, in particular transforming growth factor-beta (TGF-beta). The released factors then stimulate tumor cell signaling, which causes breast cancer cells to make increased amounts of osteolytic factors, such as parathyroid hormone-related protein (PTHrP), interleukin-11 (IL-11), and vascular endothelial growth factor (VEGF). Therefore, tumor cell-bone cell interactions cause a vicious cycle in which tumor cells stimulate bone cells to cause bone destruction. As a consequence, the local microenvironment is enriched with factors that fuel tumor growth in bone. Transforming growth factor-beta is of particular importance because it increases breast cancer production of PTHrP. Parathyroid hormone-related protein then stimulates osteoblasts to express RANK (receptor activator of nuclear factor kappa B) ligand, which in turns enhances osteoclast formation and activity. Breast cancer osteolytic metastasis can be interrupted at four points in the vicious cycle: by neutralizing PTHrP biologic activity, by blocking the TGF-beta signaling pathway in the tumor cells, by inhibiting PTHrP gene transcription, and by inhibiting bone resorption.

Topics: Animals; Bone Neoplasms; Breast Neoplasms; Female; Gene Expression Regulation, Neoplastic; Humans; Osteolysis; Parathyroid Hormone-Related Protein; Signal Transduction; Transforming Growth Factor beta

Breast carcinoma commonly metastasizes to the skeleton in patients with advanced disease to cause bone destruction and the associated pain, hypercalcemia, fracture, and nerve-compression syndromes. In this scenario, the bone destruction is mediated by the osteoclast. Tumor-produced parathyroid hormone-related protein (PTHrP), a known stimulator of osteoclastic bone resorption, is a major mediator of the osteolytic process. Transforming growth factor beta (TGFbeta), which is abundant in bone matrix and is released as a consequence of osteoclastic bone resorption, may promote breast carcinoma osteolysis by stimulating PTHrP production by tumor cells.. Stable breast carcinoma MDA-MB-231 cell lines were constructed that expressed mutant TGFbeta receptors, Smad proteins, or estrogen receptor (ER)-alpha and were used to determine the role of TGFbeta in modulating tumor production of PTHrP. These stable cell lines were applied to a mouse model of human breast carcinoma metastases to the bone to dissect the molecular mechanisms responsible for osteolytic bone metastases.. TGFbeta promoted the development and progression of osteolytic bone metastases by inducing tumor production of PTHrP, the effect of which was mediated through the Smad signaling pathway. PTHrP stimulated osteoclastic bone resorption by increasing osteoblast production of the receptor activator of nuclear factor K B (RANK) ligand and decreasing osteoblast production of osteoprotegerin (OPG). A constitutively active ER-alpha mutation (Tyr537Asn), identified from a human bone metastases, when it was expressed in human breast carcinoma cells, caused increased production of PTHrP. TGFbeta significantly enhanced the ER-alpha-mediated transcriptional activity induced by ER-alpha (Tyr537Asn), and this resulted in further stimulation of PTHrP production.. These data indicate a central role for TGFbeta in the pathogenesis of osteolytic bone metastases from breast carcinoma by 1) the induction of PTHrP through the Smad signaling pathway and 2) the potentiation of ER-alpha-mediated transcription induced by a constitutively active ER-alpha. Understanding the mechanisms of osteolysis at a molecular level will generate more effective therapeutic agents for patients with this devastating complication of cancer.

Topics: Animals; Bone Neoplasms; Bone Resorption; Breast Neoplasms; DNA-Binding Proteins; Female; Humans; Mice; Osteolysis; Parathyroid Hormone-Related Protein; Proteins; Receptors, Estrogen; Smad2 Protein; Trans-Activators; Transforming Growth Factor beta

In patients with advanced disease, several cancer types frequently metastasize to the skeleton, where they cause bone destruction. Osteolytic metastases are incurable and cause pain, hypercalcemia, fracture, and nerve compression syndromes. It was proposed over a century ago that certain cancers, such as that of the breast, preferentially metastasize to the favorable microenvironment provided by bone. Bone matrix is a rich store of immobilized growth factors that are released during bone resorption. Histological analysis of osteolytic bone metastases indicates that the bone destruction is mediated by the osteoclast rather than directly by the tumor cells. These observations suggest a vicious cycle driving the formation of osteolytic metastases: tumor cells secrete factors stimulating osteoclasts through adjacent bone marrow stromal cells; osteoclastic resorption in turn releases growth factors from the bone matrix; finally, locally released growth factors activate the tumor cells. This vicious cycle model has now been confirmed at the molecular level. In particular, transforming growth factor beta (TGF3beta) is abundant in bone matrix and released as a consequence of osteoclastic bone resorption. Bone-derived TGFbeta plays an integral role in promoting the development and progression of osteolytic bone metastases by inducing tumor production of parathyroid hormone-related protein (PTHrP), a known stimulator of osteoclastic bone resorption. In breast cancer cells TGFbeta appears to stimulate PTHrP secretion by a posttranscriptional mechanism through both Smad and p38 mitogen activated protein (MAP) kinase signaling pathways. Osteolytic metastases can be suppressed in vivo by inhibition of bone resorption, blockade of TGFbeta signaling in tumor cells, and by neutralization of PTHrP. Other factors released from bone matrix may also act on tumor cells in bone, which in turn may produce other factors that stimulate bone resorption, following the vicious cycle paradigm established for TGFbeta and PTHrP. An understanding at the molecular level of the mechanisms of osteolytic metastasis will result in more effective therapies for this devastating complication of cancer.

Topics: Bone and Bones; Bone Neoplasms; Breast Neoplasms; Calcium; Gonadal Steroid Hormones; Humans; Lymphocytes; Neovascularization, Pathologic; Osteoclasts; Osteolysis; Parathyroid Hormone-Related Protein; Proteins; Transforming Growth Factor beta

Bone appears to act like fertilizer for many tumors, including myeloma and metastatic breast cancer. The explanation must lie in interactions between tumor cells and the bone-tissue microenvironment. At this level, too, must lie the explanation of how bisphosphonates address not only cancer osteolysis but also the tumor burden. By inhibiting osteoclasts, the drugs may block a cancer-related vicious cycle.

Topics: Antineoplastic Agents; Clodronic Acid; Diphosphonates; Etidronic Acid; Humans; Neoplasm Proteins; Neoplasms; Osteolysis; Pamidronate; Parathyroid Hormone-Related Protein; Proteins; Risedronic Acid; Transforming Growth Factor beta

Topics: Animals; Breast Neoplasms; Cell Line, Tumor; Disease Models, Animal; Female; Fibronectins; HSP90 Heat-Shock Proteins; Humans; Hyaluronan Receptors; Mammary Neoplasms, Experimental; Mice; Microfilament Proteins; Osteoblasts; Osteoclasts; Osteogenesis; Osteolysis; Proteome; Secretome; Transforming Growth Factor beta; Tumor Suppressor Proteins; Wnt Signaling Pathway

While tumoral Smad-mediated transforming growth factor β (TGFβ) signaling drives osteolytic estrogen receptor α-negative (ER-) breast cancer bone metastases (BMETs) in preclinical models, its role in ER+ BMETs, representing the majority of clinical BMETs, has not been documented. Experiments were undertaken to examine Smad-mediated TGFβ signaling in human ER+ cells and bone-tropic behavior following intracardiac inoculation of estrogen (E

Topics: Animals; Apoptosis; Bone Neoplasms; Breast Neoplasms; Cell Proliferation; Female; Gene Expression Regulation, Neoplastic; Humans; Mice; Mice, Nude; Osteoclasts; Osteolysis; Receptors, Estrogen; Transforming Growth Factor beta; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

As an approach to enhance autogenous bone grafting, the fusion rate of recombinant human bone morphogenetic protein-2 (rhBMP-2) is close to 100%, which is significantly higher than other bone graft methods. However, there are some obvious problems in applying rhBMP-2 clinically. Among them, early endplate osteolysis frequently occurs in the lumbar interbody fusion, which readily leads to cage subsidence or shift, thus influencing clinical effects. Moreover, robust bone formation activity and serious osteolysis coexist. What is the internal mechanism? How do we solve this problem? Strontium (Sr) is now widely used for the treatment of osteoporosis. It elicits a double effect in that it simultaneously enhances bone formation and inhibits bone resorption. We propose that Sr might be a solution for osteolysis induced by rhBMP-2 in spinal interbody fusion. Whether this synergistic effect leads to new metabolic pathway activation remains to be explored. Clarifying the synergistic effect and mechanism will be of great importance in improving both the osteogenic effect and reducing the dose amount of rhBMP-2, as well as corresponding costs.

Topics: Bone Morphogenetic Protein 2; Humans; Lumbar Vertebrae; Osteolysis; Recombinant Proteins; Spinal Fusion; Strontium; Transforming Growth Factor beta

Multiple myeloma is a plasma cell malignancy, which develops in the bone marrow and frequently leads to severe bone destruction. Current antiresorptive therapies to treat the bone disease do little to repair damaged bone; therefore, new treatment strategies incorporating bone anabolic therapies are urgently required. We hypothesized that combination therapy using the standard of care antiresorptive zoledronic acid (Zol) with a bone anabolic (anti-TGFβ/1D11) would be more effective at treating myeloma-induced bone disease than Zol therapy alone. JJN3 myeloma-bearing mice (n = 8/group) treated with combined Zol and 1D11 resulted in a 48% increase (p ≤ 0.001) in trabecular bone volume (BV/TV) compared with Zol alone and a 65% increase (p ≤ 0.0001) compared with 1D11 alone. Our most significant finding was the substantial repair of U266-induced osteolytic bone lesions with combination therapy (n = 8/group), which resulted in a significant reduction in lesion area compared with vehicle (p ≤ 0.01) or Zol alone (p ≤ 0.01). These results demonstrate that combined antiresorptive and bone anabolic therapy is significantly more effective at preventing myeloma-induced bone disease than Zol alone. Furthermore, we demonstrate that combined therapy is able to repair established myelomatous bone lesions. This is a highly translational strategy that could significantly improve bone outcomes and quality of life for patients with myeloma. © 2018 The Authors. Journal of Bone and Mineral Research Published by Wiley Periodicals Inc.

Topics: Animals; Antibodies, Monoclonal; Cell Line, Tumor; Humans; Mice; Mice, Inbred NOD; Mice, SCID; Multiple Myeloma; Neoplasms, Experimental; Osteolysis; Transforming Growth Factor beta; Xenograft Model Antitumor Assays; Zoledronic Acid

Bone metastasis is common in renal cell carcinoma (RCC), and the lesions are mainly osteolytic. The mechanism of bone destruction in RCC bone metastasis is unknown.. We used a direct intrafemur injection of mice with bone-derived 786-O RCC cells (Bo-786) as an in vivo model to study if inhibition of osteoblast differentiation is involved in osteolytic bone lesions in RCC bone metastasis.. We showed that bone-derived Bo-786 cells induced osteolytic bone lesions in the femur of mice. We examined the effect of conditioned medium of Bo-786 cells (Bo-786 CM) on both primary mouse osteoblasts and MC3T3-E1 preosteoblasts and found that Bo-786 CM inhibited osteoblast differentiation. Secretome analysis of Bo-786 CM revealed that BIGH3 (Beta ig h3 protein), also known as TGFBI (transforming growth factor beta-induced protein), is highly expressed. We generated recombinant BIGH3 and found that BIGH3 inhibited osteoblast differentiation in vitro. In addition, CM from Bo-786 BIGH3 knockdown cells (786-BIGH3 KD) reduced the inhibition of osteoblast differentiation compared to CM from vector control. Intrafemural injection of mice with 786-BIGH3 KD cells showed a reduction in osteolytic bone lesions compared to vector control. Immunohistochemical staining of 18 bone metastasis specimens from human RCC showed strong BIGH3 expression in 11/18 (61%) and moderate BIGH3 expression in 7/18 (39%) of the specimens.. These results suggest that suppression of osteoblast differentiation by BIGH3 is one of the mechanisms that enhance osteolytic lesions in RCC bone metastasis, and raise the possibilty that treatments that increase bone formation may improve therapy outcomes.

Topics: Animals; Bone and Bones; Bone Neoplasms; Carcinoma, Renal Cell; Cell Differentiation; Cell Line, Tumor; Cell Proliferation; Disease Models, Animal; Extracellular Matrix Proteins; Gene Expression; Gene Knockout Techniques; Heterografts; Humans; Kidney Neoplasms; Mass Spectrometry; Mice; Osteoblasts; Osteolysis; Transforming Growth Factor beta; X-Ray Microtomography

The chronic inflammatory immune response triggered by the infection of the tooth root canal system results in the local upregulation of RANKL, resulting in periapical bone loss. While RANKL has a well-characterized role in the control of bone homeostasis/pathology, it can play important roles in the regulation of the immune system, although its possible immunoregulatory role in infectious inflammatory osteolytic conditions remains largely unknown. Here, we used a mouse model of infectious inflammatory periapical lesions subjected to continuous or transitory anti-RANKL inhibition, followed by the analysis of lesion outcome and multiple host response parameters. Anti-RANKL administration resulted in arrest of bone loss but interfered in the natural immunoregulation of the lesions observed in the untreated group. RANKL inhibition resulted in an unremitting proinflammatory response, persistent high proinflammatory and effector CD4 response, decreased regulatory T-cell (Treg) migration, and lower levels of Treg-related cytokines IL-10 and TGFb. Anti-RANKL blockade impaired the immunoregulatory process only in early disease stages, while the late administration of anti-RANKL did not interfere with the stablished immunoregulation. The impaired immunoregulation due to RANKL inhibition is characterized by increased delayed-type hypersensitivity in vivo and T-cell proliferation in vitro to the infecting bacteria, which mimic the effects of Treg inhibition, reinforcing a possible influence of RANKL on Treg-mediated suppressive response. The adoptive transfer of CD4+FOXp3+ Tregs to mice receiving anti-RANKL therapy restored the immunoregulatory capacity, attenuating the inflammatory response in the lesions, reestablishing normal T-cell response in vivo and in vitro, and preventing lesion relapse upon anti-RANKL therapy cessation. Therefore, while RANKL inhibition efficiently limited the periapical bone loss, it promoted an unremitting host inflammatory response by interfering with Treg activity, suggesting that this classic osteoclastogenic mediator plays a role in immunoregulation.

Topics: Adoptive Transfer; Alveolar Bone Loss; Animals; Antibodies, Monoclonal; Cell Proliferation; Cell Survival; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Gene Expression; Immunity, Mucosal; Inflammation; Infliximab; Interleukin-10; Lymphocyte Activation; Male; Mice; Mice, Inbred C57BL; Osteolysis; Periapical Diseases; RANK Ligand; Real-Time Polymerase Chain Reaction; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

High recurrence and lower survival rates in patients with oral squamous cell carcinoma (OSCC) are associated with its bone invasion. We identified the oncogenic role of RUNX3 during bone invasion by OSCC. Tumor growth and the generation of osteolytic lesions were significantly inhibited in mice that were subcutaneously inoculated with RUNX3-knockdown human OSCC cells. RUNX3 knockdown enhanced TGF-β-induced growth arrest and inhibited OSCC cell migration and invasion in the absence or presence of transforming growth factor-β (TGF-β), a major growth factor abundant in the bone microenvironment. RUNX3 knockdown induced cell cycle arrest at the G1 and G2 phases and promoted G2 arrest by TGF-β in Ca9.22 OSCC cells. RUNX3 knockdown also inhibited both the basal and TGF-β-induced epithelial-to-mesenchymal transition by increasing E-cadherin expression and suppressing the nuclear translocation of β-catenin. In addition, the expression and TGF-β-mediated induction of parathyroid hormone-related protein (PTHrP), one of key osteolytic factors, was blocked in RUNX3-knockdown OSCC cells. Furthermore, treating human osteoblastic cells with conditioned medium derived from RUNX3-knockdown OSCC cells reduced the receptor activator of nuclear factor-kappaB ligand (RANKL)/osteoprotegerin ratio compared with treatment with conditioned medium from RUNX3-expressing cells. These findings indicate that RUNX3 expression in OSCC cells contributes to their bone invasion and the resulting osteolysis by inducing their malignant behaviors and production of osteolytic factors. RUNX3 alone or in combination with TGF-β and PTHrP may be a useful predictive biomarker and therapeutic target for bone invasion by oral cancer.

Topics: Animals; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Movement; Cell Proliferation; Core Binding Factor Alpha 3 Subunit; Epithelial-Mesenchymal Transition; G1 Phase Cell Cycle Checkpoints; G2 Phase Cell Cycle Checkpoints; Gene Expression Regulation, Neoplastic; Head and Neck Neoplasms; Humans; Kaplan-Meier Estimate; Male; Mice, Inbred BALB C; Mice, Nude; Mouth Neoplasms; Neoplasm Invasiveness; Osteoblasts; Osteolysis; Paracrine Communication; Parathyroid Hormone-Related Protein; RANK Ligand; RNA Interference; Signal Transduction; Skull; Squamous Cell Carcinoma of Head and Neck; Time Factors; Transfection; Transforming Growth Factor beta; Tumor Microenvironment

The Fifth Biennial Research Summit of the American Association of Oral and Maxillofacial Surgeons and its Committee on Research Planning and Technology Assessment was held in Rosemont, Illinois on May 6 and 7, 2015. The goal of the symposium is to provide a forum for the most recent clinical and scientific advances to be brought to the specialty. The proceedings of the events of that summit are presented in this report.

Topics: Ameloblastoma; Bone Morphogenetic Protein 2; Bone Regeneration; Carcinogenesis; Congresses as Topic; Dental Research; Fibrous Dysplasia of Bone; Humans; Imaging, Three-Dimensional; Nerve Growth Factor; Odontogenic Tumors; Osteolysis; Patient Care Planning; Plastic Surgery Procedures; Printing, Three-Dimensional; Recombinant Proteins; Regenerative Medicine; Registries; Societies, Dental; Surgery, Computer-Assisted; Surgery, Oral; Tissue Engineering; Transforming Growth Factor beta; Translational Research, Biomedical; User-Computer Interface

Transforming growth factor (TGF)-β supports multiple myeloma progression and associated osteolytic bone disease. Conversion of latent TGF-β to its biologically active form is a major regulatory node controlling its activity. Thrombospondin1 (TSP1) binds and activates TGF-β. TSP1 is increased in myeloma, and TSP1-TGF-β activation inhibits osteoblast differentiation. We hypothesized that TSP1 regulates TGF-β activity in myeloma and that antagonism of the TSP1-TGF-β axis inhibits myeloma progression. Antagonists (LSKL peptide, SRI31277) derived from the LSKL sequence of latent TGF-β that block TSP1-TGF-β activation were used to determine the role of the TSP1-TGF-β pathway in mouse models of myeloma. TSP1 binds to human myeloma cells and activates TGF-β produced by cultured human and mouse myeloma cell lines. Antagonists delivered via osmotic pump in an intratibial severe combined immunodeficiency CAG myeloma model or in a systemic severe combined immunodeficiency CAG-heparanase model of aggressive myeloma reduced TGF-β signaling (phospho-Smad 2) in bone sections, tumor burden, mouse IL-6, and osteoclasts, increased osteoblast number, and inhibited bone destruction as measured by microcomputed tomography. SRI31277 reduced tumor burden in the immune competent 5TGM1 myeloma model. SRI31277 was as effective as dexamethasone or bortezomib, and SRI31277 combined with bortezomib showed greater tumor reduction than either agent alone. These studies validate TSP1-regulated TGF-β activation as a therapeutic strategy for targeted inhibition of TGF-β in myeloma.

Topics: Animals; Antineoplastic Agents; Bortezomib; Cell Differentiation; Disease Models, Animal; Humans; Interleukin-6; Male; Mice; Mice, Inbred C57BL; Mice, SCID; Multiple Myeloma; Osteogenesis; Osteolysis; Peptides; Random Allocation; Signal Transduction; Thrombospondin 1; Transforming Growth Factor beta; Tumor Microenvironment

We report on a boy who presented with hypophosphatemic rickets with elevated serum fibroblast growth factor 23 (FGF23) and polyostotic osteolytic lesions at age 2 years. Tumor-induced hypophosphatemic rickets was suspected; however, bone biopsy for osteolytic changes revealed no tumorous change, except for irregularly dilated vessels associated with osteoclasts and fibrous proliferation. Venous sampling failed to point to FGF23-producing foci. After alfacalcidol and phosphate supplementation, the rachitic skeletal changes improved, but FGF23 increased and new osteolytic lesions developed. Serum levels of neopterin and a few cytokines, including plasma transforming growth factor-β and soluble tumor necrosis factor receptor type II, were elevated. At age 4 years, high doses of phosphate resulted in increased serum phosphate levels, decreased neopterin and cytokines, decreased FGF23, and stabilization of osteolysis. We excluded germline mutations in PHEX, FGF23, DMP1, and ENPP1 (genes for hereditary hypophosphatemic rickets) and somatic mutations in the GNAS and HRAS/KRAS (the disease-causing genes for McCune-Albright syndrome and linear nevus sebaceous syndrome, respectively). We could not perform octreotide scintigraphy or fluorodeoxyglucose-positron emission tomography, and thus could not completely exclude occult FGF23-producing tumors. However, considering the course of the disease, it is intriguing to assume that dysregulation of osteoclast-macrophage lineage may have induced increased neopterin levels, increased cytokine levels, osteolytic process, and possibly FGF23 overproduction.

Topics: Cell Proliferation; Child, Preschool; Cytokines; Fibroblast Growth Factor-23; Fibroblast Growth Factors; Fibroblasts; Fibrous Dysplasia, Polyostotic; Gene Expression Regulation; Humans; Hydroxycholecalciferols; Male; Neopterin; Osteoclasts; Osteolysis; Phosphates; Receptors, Tumor Necrosis Factor, Type II; Rickets, Hypophosphatemic; Transforming Growth Factor beta

Bone metastasis affects more than 70% of advanced breast cancer patients, but the molecular mechanisms of this process remain unclear. Here, we present clinical and experimental evidence to clarify the role of the integrin β-like 1 (ITGBL1) as a key contributor to bone metastasis of breast cancer. In an in vivo model system and in vitro experiments, ITGBL1 expression promoted formation of osteomimetic breast cancers, facilitating recruitment, residence, and growth of cancer cells in bone microenvironment along with osteoclast maturation there to form osteolytic lesions. Mechanistic investigations identified the TGFβ signaling pathway as a downstream effector of ITGBL1 and the transcription factor Runx2 as an upstream activator of ITGBL1 expression. In support of these findings, we also found that ITGBL1 was an essential mediator of Runx2-induced bone metastasis of breast cancer. Overall, our results illuminate how bone metastasis occurs in breast cancer, and they provide functional evidence for new candidate biomarkers and therapeutic targets to identify risk, to prevent, and to treat this dismal feature of advanced breast cancer.

Topics: Animals; Blotting, Western; Bone Neoplasms; Breast Neoplasms; Cell Line; Cell Line, Tumor; Cell Movement; Cell Survival; Cells, Cultured; Core Binding Factor Alpha 1 Subunit; Female; Gene Expression Regulation, Neoplastic; Humans; Integrin beta1; Kaplan-Meier Estimate; Mice, Inbred BALB C; Mice, SCID; Osteoblasts; Osteogenesis; Osteolysis; Reverse Transcriptase Polymerase Chain Reaction; RNA Interference; Signal Transduction; Transforming Growth Factor beta

Cancer patients frequently develop skeletal metastases that significantly impact quality of life. Since bone metastases remain incurable, a clearer understanding of molecular mechanisms regulating skeletal metastases is required to develop new therapeutics that block establishment of tumors in bone. While many studies have suggested that the microenvironment contributes to bone metastases, the factors mediating tumors to progress from a quiescent to a bone-destructive state remain unclear. In this study, we hypothesized that the "soil" of the bone microenvironment, specifically the rigid mineralized extracellular matrix, stimulates the transition of the tumor cells to a bone-destructive phenotype. To test this hypothesis, we synthesized 2D polyurethane (PUR) films with elastic moduli ranging from the basement membrane (70 MPa) to cortical bone (3800 MPa) and measured expression of genes associated with mechanotransduction and bone metastases. We found that expression of Integrin β3 (Iβ3), as well as tumor-produced factors associated with bone destruction (Gli2 and parathyroid hormone related protein (PTHrP)), significantly increased with matrix rigidity, and that blocking Iβ3 reduced Gli2 and PTHrP expression. To identify the mechanism by which Iβ3 regulates Gli2 and PTHrP (both are also known to be regulated by TGF-β), we performed Förster resonance energy transfer (FRET) and immunoprecipitation, which indicated that Iβ3 co-localized with TGF-β Receptor Type II (TGF-β RII) on rigid but not compliant films. Finally, transplantation of tumor cells expressing Iβ3 shRNA into the tibiae of athymic nude mice significantly reduced PTHrP and Gli2 expression, as well as bone destruction, suggesting a crucial role for tumor-produced Iβ3 in disease progression. This study demonstrates that the rigid mineralized bone matrix can alter gene expression and bone destruction in an Iβ3/TGF-β-dependent manner, and suggests that Iβ3 inhibitors are a potential therapeutic approach for blocking tumor transition to a bone destructive phenotype.

Topics: Adenocarcinoma; Animals; Bone Neoplasms; Breast Neoplasms; Carcinoma, Squamous Cell; Cell Line, Tumor; Elastic Modulus; Extracellular Matrix; Female; Gene Expression Regulation, Neoplastic; Humans; Integrin beta3; Kruppel-Like Transcription Factors; Lung Neoplasms; Mice; Mice, Nude; Neoplasm Proteins; Nuclear Proteins; Osteolysis; Pliability; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; Transfection; Transforming Growth Factor beta; Tumor Microenvironment; Xenograft Model Antitumor Assays; Zinc Finger Protein Gli2

Cancer-associated muscle weakness is a poorly understood phenomenon, and there is no effective treatment. Here we find that seven different mouse models of human osteolytic bone metastases-representing breast, lung and prostate cancers, as well as multiple myeloma-exhibited impaired muscle function, implicating a role for the tumor-bone microenvironment in cancer-associated muscle weakness. We found that transforming growth factor (TGF)-β, released from the bone surface as a result of metastasis-induced bone destruction, upregulated NADPH oxidase 4 (Nox4), resulting in elevated oxidization of skeletal muscle proteins, including the ryanodine receptor and calcium (Ca(2+)) release channel (RyR1). The oxidized RyR1 channels leaked Ca(2+), resulting in lower intracellular signaling, which is required for proper muscle contraction. We found that inhibiting RyR1 leakage, TGF-β signaling, TGF-β release from bone or Nox4 activity improved muscle function in mice with MDA-MB-231 bone metastases. Humans with breast- or lung cancer-associated bone metastases also had oxidized skeletal muscle RyR1 that is not seen in normal muscle. Similarly, skeletal muscle weakness, increased Nox4 binding to RyR1 and oxidation of RyR1 were present in a mouse model of Camurati-Engelmann disease, a nonmalignant metabolic bone disorder associated with increased TGF-β activity. Thus, pathological TGF-β release from bone contributes to muscle weakness by decreasing Ca(2+)-induced muscle force production.

Topics: Absorptiometry, Photon; Animals; Bone Neoplasms; Breast Neoplasms; Calcium; Calcium Signaling; Camurati-Engelmann Syndrome; Cell Line, Tumor; Disease Models, Animal; Female; Humans; Lung Neoplasms; Male; MCF-7 Cells; Mice; Mice, Nude; Mice, SCID; Multiple Myeloma; Muscle Contraction; Muscle Proteins; Muscle Strength; Muscle Weakness; Muscle, Skeletal; NADPH Oxidase 4; NADPH Oxidases; Neoplasms; Osteolysis; Oxidation-Reduction; Prostatic Neoplasms; Ryanodine Receptor Calcium Release Channel; Transforming Growth Factor beta; Up-Regulation; X-Ray Microtomography

Since the introduction of rhBMP-2 (Infuse) in 2002, surgeons have had an alternative substitute to autograft and its related donor site morbidity. Recently, the prevalence of reported adverse events and complications related to the use of rhBMP-2 has raised many ethical and legal concerns for surgeons. Additionally, the cost and decreasing reimbursement landscape of rhBMP-2 use have required identification of a viable alternative. Osteo allogeneic morphogenetic protein (OsteoAMP) is a commercially available allograft-derived growth factor rich in osteoinductive, angiogenic, and mitogenic proteins. This study compares the radiographic fusion outcomes between rhBMP-2 and OsteoAMP allogeneic morphogenetic protein in lumbar interbody fusion spine procedures.. Three hundred twenty-one (321) patients from three centers underwent a transforaminal lumbar interbody fusion (TLIF) or lateral lumbar interbody fusion (LLIF) procedure and were assessed by an independent radiologist for fusion and radiographically evident complications. The independent radiologist was blinded to the intervention, product, and surgeon information. Two hundred and twenty-six (226) patients received OsteoAMP with autologous local bone, while ninety-five (95) patients received Infuse with autologous local bone. Patients underwent radiographs (x-ray and/or CT) at standard postoperative follow-up intervals of approximately 1, 3, 6, 12, and 18 months. Fusion was defined as radiographic evidence of bridging across endplates, or bridging from endplates to interspace disc plugs. Osteobiologic surgical supply costs were also analyzed to ascertain cost differences between OsteoAMP and rhBMP-2.. OsteoAMP produced higher rates of fusion at 6, 12, and 18 months (p ≤ 0.01). The time required for OsteoAMP to achieve fusion was approximately 40% less than rhBMP-2 with approximately 70% fewer complications. Osteobiologic supply costs were 80.5% lower for OsteoAMP patients (73.7% lower per level) than for rhBMP-2.. Results of this study indicate that OsteoAMP is a viable alternative to rhBMP-2 both clinically and economically when used in TLIF and LLIF spine procedures.

Topics: Adult; Aged; Bone Morphogenetic Protein 2; Bone Morphogenetic Proteins; Bone Transplantation; Drug Costs; Drug Evaluation; Female; Humans; Lumbar Vertebrae; Male; Middle Aged; Ossification, Heterotopic; Osteolysis; Radiography; Recombinant Proteins; Retrospective Studies; Single-Blind Method; Spinal Fusion; Transforming Growth Factor beta; Treatment Outcome

Effects of curcuminoids on breast cancer cell secretion of the bone-resorptive peptide parathyroid hormone-related protein (PTHrP) and on lytic breast cancer bone metastasis were evaluated. In vitro, transforming growth factor (TGF)-β-stimulated PTHrP secretion was inhibited by curcuminoids (IC50 = 24 μM) in MDA-MB-231 human breast cancer cells independent of effects on cell growth inhibition. Effects on TGF-β signaling revealed decreases in phospho-Smad2/3 and Ets-1 protein levels with no effect on p-38 MAPK-mediated TGF-β signaling. In vivo, mice were inoculated with MDA-MB-231 cells into the left cardiac ventricle and treated ip every other day with curcuminoids (25 or 50 mg/kg) for 21 days. Osteolytic bone lesion area was reduced up to 51% (p < 0.01). Consistent with specific effects on bone osteolysis, osteoclast number at the bone-tumor interface was reduced up to 53% (p < 0.05), while tumor area within bone was unaltered. In a separate study, tumor mass in orthotopic mammary xenografts was also unaltered by treatment. These data suggest that curcuminoids prevent TGF-β induction of PTHrP and reduce osteolytic bone destruction by blockade of Smad signaling in breast cancer cells.

Topics: Animals; Bone Neoplasms; Breast Neoplasms; Curcumin; Disease Models, Animal; Female; Humans; Mice; Molecular Structure; Osteolysis; Parathyroid Hormone-Related Protein; Signal Transduction; Transforming Growth Factor beta

Metastasis is the rapid proliferation of cancer cells (secondary tumour) at a specific place, generally leading to death. This occurs at anatomical parts providing the necessary environment for vascularity, oxygen and food to hide their actions and trigger the rapid growth of cancer. Prostate and breast cancers, for example, use bone marrow for their proliferation. Bone-supporting cancer cells thus adapt to the environment, mimicking the behaviour of genetic and molecular bone cells. Evidence of this has been given in Cecchini et al. (2005, EAU Update Ser. 3:214-226), providing arguments such as how cancer cell growth is so active during bone reabsorption. This paper simulates metastasis activation in bone marrow. A mathematical model has been developed involving the activation of molecules from bone tissue cells, which are necessary for cancer to proliferate. Here, we simulate two forms of secondary tumour growth depending on the type of metastasis: osteosclerosis and osteolysis.

Topics: Biomedical Engineering; Bone Marrow Neoplasms; Bone Neoplasms; Bone Remodeling; Cell Differentiation; Cell Proliferation; Computer Simulation; Humans; Mathematical Concepts; Models, Biological; Neoplasm Metastasis; Osteolysis; Osteosclerosis; Parathyroid Hormone-Related Protein; Somatomedins; Transforming Growth Factor beta

To observe the effect of a slow-release recombinant human bone morphogenetic protein-2 (rhBMP-2) formulation on the expressions of receptor activator of nuclear factor-κB ligand (RANKL) and osteoprotegerin (OPG) in a murine air pouch model of bone implantation.. A cranial bone allograft was implanted in the air pouch induced on the back of the recipients. The rat models were then randomized into 5 groups, including a blank control group, chromium particle group, and 3 rhBMP-2 groups receiving 50, 100 or 200 µg/L slow-release rhBMP-2 in addition to chromium particles. Three weeks later, the expressions of RANKL and OPG in the air pouch was detected using Western blotting and RT-PCR, and the positively stained area for osteoclasts in the bone graft was determined with TRAP staining for drug effect assessment.. RANKL and OPG expressions were found in the air pouches in all the 5 groups. RANKL and OPG protein and mRNA expressions, RANKL/OPG ratio and osteoclast staining area in the bone graft were the highest in chromium particle group (P<0.05), but were significantly decreased by treatment with the slow-release rhBMP-2 formulation (P<0.05); the measurements showed no significant differences between the blank control group and 200 µg/L rhBMP-2 group (P>0.05).. Chromium particles can cause osteolysis by increasing the RANKL/OPG ratio in rats, and intervention with slow-release rhBMP-2 can significantly promote bone formation and suppress bone resorption by decreasing RANKL/OPG ratio.

Topics: Animals; Arthroplasty, Replacement; Bone Morphogenetic Protein 2; Bone Resorption; Bone Transplantation; Chromium; Delayed-Action Preparations; Disease Models, Animal; Female; Male; Osteolysis; Particle Size; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Skull; Transforming Growth Factor beta

Langerhans cell histiocytosis (LCH) has a challenging and still unclear pathogenesis. A body of literature points to impaired maturation of the lesional dendritic cells, and to immune dysregulation in the form of increased FoxP3 cells. Various cytokine abnormalities such as expression of transforming growth factor (TGF)-β have been reported, as well as abnormalities in lipid content in LCH cells. Morphoproteomic techniques were applied to identify the signal transduction pathways that could influence histogenesis and immune regulation in osteolytic LCH. Five pediatric cases of osteolytic LCH were examined, using antibodies against CD1a, S100, CD68, CD8, FoxP3, phosphorylated (p)-STAT3 (Tyr705), protein kinase C (PKC)-α, phospholipase (PL)D1, fatty acid synthase (FASN), and zinc finger protein, Gli2. Positive and negative controls were performed. A FoxP3(+)/CD8(+) cell ratio was calculated by counting the FoxP3+ and CD8+ cells in 10 high power fields for each case. There is induction of sonic hedgehog (SHH) mediators consistent with TGF-β signaling pathway through Smad3-dependent activation of Gli2, findings supported by the plasmalemmal and cytoplasmic expression of PKC-α and PLD1, and nuclear expression of Gli2, in lesional cells. The FoxP3+/CD8+ cell ratio is increased, ranging from 1.7-7.94. There is moderate cytoplasmic expression of FASN in most of the Langerhans cells, a finding that supports previously published phospholipid abnormalities in LCH and is consistent with PKC-α/PLD1/TGF-β signaling. With our study, we strongly suggest that the TGF-β cell signaling pathway is a major player in the pathogenesis of LCH, leading to non-canonical induction of nuclear Gli2 expression, thereby contributing to osteoclastogenesis in LCH histiocytes. It could also cause a state of immune frustration in LCH, by inducing the transformation of CD4(+)CD25(-) cells into CD4(+)/FoxP3(+) cells. This coincides with the clinical evidence of a response to thalidomide in patients with osteolytic LCH, given its reported ability to reduce TGF-beta 1 and FoxP3 cells. Such TGF-β signaling in osteoclastogenesis and immune dysregulation, and the presence of FASN in the majority of cells, have additional therapeutic implications for osteolytic LCH.

Topics: Biomarkers; Cell Differentiation; Cell Membrane; Cell Nucleus; Child; Child, Preschool; Cytoplasm; Female; Histiocytosis, Langerhans-Cell; Humans; Kruppel-Like Transcription Factors; Male; Nuclear Proteins; Osteoclasts; Osteolysis; Proteomics; Signal Transduction; Thalidomide; Transforming Growth Factor beta; Zinc Finger Protein Gli2

Using bone morphogenic protein (BMP) to augment fusion in spine surgery is widespread and lends itself in particular to minimally invasive lumbar fusion, where the surface area for fusion is significantly less than the equivalent open procedure.. Here we described the use of very low-dose BMP in promoting fusion in minimally invasive lumbar interbody fixation but also highlight some of the potential complications of BMP-2 use and techniques available to reduce or avoid them.. Prospective observational study of consecutive patients undergoing minimally invasive lumbar interbody fusion with percutaneous pedicle screws.. Thirty patients aged between 22 and 78 years (mean 53 years).. Thin-slice lumbar computed tomography scanning with multiplanar reconstruction at 6 and 12 months postoperative.. Thirty-six spinal levels were instrumented in total, of which four underwent posterior lumbar interbody fusion and 32 underwent transforaminal lumbar interbody fusion. Bone graft harvested locally was placed in the disc space with low-dose BMP-2 (1.4 mg per level).. Thirty-three of 36 spinal levels showed complete fusion at a mean postoperative scan time of 7.1 months. Two levels demonstrated partial fusion at 6 months, which was complete at 12 months. There was one case of nonunion at 12 months, which also demonstrated vertebral body osteolysis. Despite very low-dose BMP-2, two cases of asymptomatic heterotopic ossification were observed, and there were two cases of perineural cyst formation, one of whom required revision of the interbody cage.. The use of BMP with autograft in the disc space during minimally invasive lumbar interbody fusion is associated with a high rate of early fusion. Even with very low-dose BMP used in this study, complications related to BMP usage were not avoided completely.

Topics: Adult; Aged; Bone Morphogenetic Protein 2; Cysts; Female; Humans; Lumbar Vertebrae; Male; Middle Aged; Ossification, Heterotopic; Osteolysis; Postoperative Complications; Recombinant Proteins; Spinal Fusion; Transforming Growth Factor beta; Treatment Outcome; Young Adult

Recent studies have demonstrated cases of adjacent vertebral body osteolysis when assessing the effect of bone morphogenetic protein (BMP) on fusion rates. However, no study to date has evaluated the course of osteolysis at different periods.. To determine the incidence and resolution of osteolysis associated with BMP used in transforaminal lumbar interbody fusions (TLIF).. Retrospective review.. All TLIF cases using BMP performed at one institution with routine postoperative computed tomography (CT) scans at defined intervals.. Area of osteolysis and fusion as determined by CT scan.. We performed a retrospective analysis of all patients at our facility who underwent TLIF with BMP. Included were all patients who had obtained a CT scan within 48 hours of surgery, 3 to 6 months postoperatively, and 1 to 2 years postoperatively. Areas of osteolysis were defined as lucency within the vertebral body communicating with the interbody spacer that was not present on the immediately postoperative CT scan. Areas of osteolysis were measured in all three planes and the volume used for comparison of the 3 to 6 months CT scans with the greater than 1 year CT scan.. Twenty-three patients who underwent TLIF with BMP had obtained CT scans at all time periods required for evaluation. Seventy-eight vertebral bodies/end plates were assessed for osteolysis (39 levels). The incidence of osteolysis 3 to 6 months postoperatively in the adjacent vertebral bodies was 54% compared with 41% at 1 to 2 years. The mean volume of osteolysis was at 0.216 cm(3) at 1 to 2 years compared with 0.306 cm(3) at 3 to 6 months (p=.082). The area/rate of osteolysis did not appear to significantly affect the rate of fusion or final outcome with an overall union rate of 83%.. The rate of osteolysis decreased at 1 year compared with 3 to 6 months, but only 24% of the vertebral bodies with evidence of osteolysis at 3 to 6 months completely resolved by 1 year.

Topics: Adult; Aged; Aged, 80 and over; Bone Morphogenetic Protein 2; Female; Humans; Incidence; Lumbar Vertebrae; Male; Middle Aged; Osteolysis; Postoperative Complications; Recombinant Proteins; Retrospective Studies; Spinal Fusion; Tomography, X-Ray Computed; Transforming Growth Factor beta; Young Adult

Topics: Bone Morphogenetic Protein 2; Female; Humans; Middle Aged; Osteolysis; Postoperative Complications; Recombinant Proteins; Spinal Fusion; Spine; Surgical Sponges; Tomography, X-Ray Computed; Transforming Growth Factor beta

Breast cancer is the second leading cause of cancer-related death in women in the United States. During the advanced stages of disease, many breast cancer patients suffer from bone metastasis. These metastases are predominantly osteolytic and develop when tumor cells interact with bone. In vivo models that mimic the breast cancer-specific osteolytic bone microenvironment are limited. Previously, we developed a mouse model of tumor-bone interaction in which three mouse breast cancer cell lines were implanted onto the calvaria. Analysis of tumors from this model revealed that they exhibited strong bone resorption, induction of osteoclasts and intracranial penetration at the tumor bone (TB)-interface.. In this study, we identified and used a TB microenvironment-specific gene expression signature from this model to extend our understanding of the metastatic bone microenvironment in human disease and to predict potential therapeutic targets.. We identified a TB signature consisting of 934 genes that were commonly (among our 3 cell lines) and specifically (as compared to tumor-alone area within the bone microenvironment) up- and down-regulated >2-fold at the TB interface in our mouse osteolytic model. By comparing the TB signature with gene expression profiles from human breast metastases and an in vitro osteoclast model, we demonstrate that our model mimics both the human breast cancer bone microenvironment and osteoclastogenesis. Furthermore, we observed enrichment in various signaling pathways specific to the TB interface; that is, TGF-β and myeloid self-renewal pathways were activated and the Wnt pathway was inactivated. Lastly, we used the TB-signature to predict cyclopenthiazide as a potential inhibitor of the TB interface.. Our mouse breast cancer model morphologically and genetically resembles the osteoclastic bone microenvironment observed in human disease. Characterization of the gene expression signature specific to the TB interface in our model revealed signaling mechanisms operative in human breast cancer metastases and predicted a therapeutic inhibitor of cancer-mediated osteolysis.

Topics: Animals; Bone Neoplasms; Cell Line, Tumor; Cluster Analysis; Female; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Histocytochemistry; Humans; Mammary Neoplasms, Experimental; Mice; Osteoclasts; Osteolysis; Reproducibility of Results; Signal Transduction; Transforming Growth Factor beta; Tumor Microenvironment; Wnt Proteins

The tropism of breast cancer cells for bone and their tendency to induce an osteolytic phenotype are a result of interactions between breast cancer cells and stromal cells and are of paramount importance for bone metastasis. However, the underlying molecular mechanisms remain poorly understood. We hypothesize that tumor-stromal interaction alters gene expression in malignant tumor cells and stromal cells creating a unique expression signature that promotes osteolytic breast cancer bone metastasis and that inhibition of such interactions can be developed as targeted therapeutics. Microarray analysis was performed to investigate gene expression profiling at the tumor-bone (TB) interface versus the tumor alone area from syngenic mice injected with three different syngenic mammary tumor cell lines that differ in their metastatic potential. We identified matrix metalloproteinase 13 (MMP13), receptor activator of NF-kappaB ligand (RANKL), and integrins binding sialoprotein to be genes upregulated at the TB interface and validated. To determine the functional role of MMP13 in tumor-induced osteolysis, mice with Cl66 mammary tumors were treated with MMP13 antisense oligonucleotides (MMP13-ASO) or control scrambled oligonucleotides (control-ASO). Knockdown of MMP13 expression at the TB interface leads to significant reduction in bone destruction and in the number of activated osteoclasts at the TB interface. Further analysis to evaluate the mechanism of MMP13-dependent osteolytic bone metastasis revealed that MMP13-ASO treatment decreased active MMP9, RANKL levels, and transforming growth factor-beta signaling at the TB interface. Together, our data indicate that upregulation of MMP13 at the TB interface is important in tumor-induced osteolysis and suggest that MMP13 is a potential therapeutic target for breast cancer bone metastasis.

Topics: Adenocarcinoma; Animals; Bone and Bones; Bone Neoplasms; Cell Line, Tumor; Enzyme Activation; Female; Gene Expression Profiling; Mammary Neoplasms, Experimental; Matrix Metalloproteinase 13; Matrix Metalloproteinase 9; Matrix Metalloproteinase Inhibitors; Mice; Mice, Inbred BALB C; Osteoclasts; Osteolysis; Osteoprotegerin; RANK Ligand; RNA, Messenger; Signal Transduction; Transforming Growth Factor beta; Up-Regulation

cAMP-response-element-binding protein (CREB) signaling has been reported to be associated with cancer development and poor clinical outcome in various types of cancer. However, it remains to be elucidated whether CREB is involved in breast cancer development and osteotropism. Here, we found that metastatic MDA-MB-231 breast cancer cells exhibited higher CREB expression than did non-metastatic MCF-7 cells and that CREB expression was further increased by several soluble factors linked to cancer progression, such as IL-1, IGF-1, and TGF-beta. Using wild-type CREB and a dominant-negative form (K-CREB), we found that CREB signaling positively regulated the proliferation, migration, and invasion of MDA-MB-231 cells. In addition, K-CREB prevented MDA-MB-231 cell-induced osteolytic lesions in a mouse model of cancer metastasis. Furthermore, CREB signaling in cancer cells regulated the gene expression of PTHrP, MMPs, and OPG, which are closely involved in cancer metastasis and bone destruction. These results indicate that breast cancer cells acquire CREB overexpression during their development and that this CREB upregulation plays an important role in multiple steps of breast cancer bone metastasis.

Topics: Animals; Bone Neoplasms; Breast Neoplasms; Cell Line, Tumor; Cyclic AMP Response Element-Binding Protein; Female; Gene Expression Regulation, Neoplastic; Humans; Insulin-Like Growth Factor I; Interleukin-17; Matrix Metalloproteinases; Mice; Mice, Nude; Neoplasm Metastasis; Osteolysis; Osteoprotegerin; Parathyroid Hormone-Related Protein; Transforming Growth Factor beta; Xenograft Model Antitumor Assays

Gingival squamous cell carcinoma (SCC) cells frequently invade mandibular bone, and this destruction is associated with a worse prognosis. However, the relationship between bone destruction and associated factors is unclear. In this study, the role and diagnostic utility of transforming growth factor-beta (TGF-beta) type I receptor (TbetaRI) in bone destruction of the mandible was investigated.. The expression of TbetaRI was explored by using an immunohistochemical method on paraffin-embedded tissues from 21 cases of mandibular SCC. An inhibitor of the kinase activity of the TbetaRI (TbetaRI-I) was used to assess the role of TbetaRI in bone destruction by a human oral SCC cell line (HSC-2) that highly expresses TbetaRI.. TbetaRI-positive signals were closely associated with destructive invasion of the mandible by oral SCC cells. Consistent with these results, TbetaRI-I greatly reduced HSC-2 cell-induced bone destruction and osteoclast formation in vivo and in vitro. TbetaRI-I treatment reduced the expression of TNF-alpha, RANKL and connective tissue growth factor (CTGF/CCN2), all of which were up-regulated by TGF-beta in HSC-2 cells.. These data demonstrated an important role for TGF-beta signalling in bone invasion by oral SCC cells, and suggest that the bone destruction is mediated by RANKL, TNF-alpha and CCN2.

Topics: Aged; Aged, 80 and over; Animals; Carcinoma, Squamous Cell; Cell Line, Tumor; Connective Tissue Growth Factor; Female; Gene Expression; Gingival Neoplasms; Humans; Male; Mice; Mice, Nude; Middle Aged; Osteolysis; Protein Serine-Threonine Kinases; RANK Ligand; Receptor, Transforming Growth Factor-beta Type I; Receptors, Transforming Growth Factor beta; Signal Transduction; Transforming Growth Factor beta; Transplantation, Heterologous; Tumor Necrosis Factor-alpha

Nearly 70% of breast cancer patients with advanced disease will develop bone metastases. Once established in bone, tumor cells produce factors that cause changes in normal bone remodeling, such as parathyroid hormone-related protein (PTHrP). While enhanced expression of PTHrP is known to stimulate osteoclasts to resorb bone, the environmental factors driving tumor cells to express PTHrP in the early stages of development of metastatic bone disease are unknown. In this study, we have shown that tumor cells known to metastasize to bone respond to 2D substrates with rigidities comparable to that of the bone microenvironment by increasing expression and production of PTHrP. The cellular response is regulated by Rho-dependent actomyosin contractility mediated by TGF-ß signaling. Inhibition of Rho-associated kinase (ROCK) using both pharmacological and genetic approaches decreased PTHrP expression. Furthermore, cells expressing a dominant negative form of the TGF-ß receptor did not respond to substrate rigidity, and inhibition of ROCK decreased PTHrP expression induced by exogenous TGF-ß. These observations suggest a role for the differential rigidity of the mineralized bone microenvironment in early stages of tumor-induced osteolysis, which is especially important in metastatic cancer since many cancers (such as those of the breast and lung) preferentially metastasize to bone.

Topics: Algorithms; Bone Neoplasms; Breast Neoplasms; Cell Culture Techniques; Cell Line, Tumor; Female; Gene Expression Regulation, Neoplastic; Gene Regulatory Networks; Humans; Neoplasm Metastasis; Osteoclasts; Osteolysis; Parathyroid Hormone-Related Protein; Reverse Transcriptase Polymerase Chain Reaction; rho-Associated Kinases; Transforming Growth Factor beta; Tumor Microenvironment

Understanding the cellular and molecular changes in the bone microenvironment is important for developing novel therapeutics to control breast cancer bone metastasis. Although the underlying mechanism(s) of bone metastasis has been the focus of intense investigation, relatively little is known about complex molecular interactions between malignant cells and bone stroma. Using a murine syngeneic model that mimics osteolytic changes associated with human breast cancer, we examined the role of tumor-bone interaction in tumor-induced osteolysis and malignant growth in the bone microenvironment. We identified transforming growth factor-beta receptor 1 (TGF-betaRI) as a commonly upregulated gene at the tumor-bone (TB) interface. Moreover, TGF-betaRI expression and activation, analyzed by nuclear localization of phospho-Smad2, was higher in tumor cells and osteoclasts at the TB interface as compared to the tumor-alone area. Furthermore, attenuation of TGF-beta activity by neutralizing antibody to TGF-beta or TGF-betaRI kinase inhibitor reduced mammary tumor-induced osteolysis, TGF-betaRI expression and its activation. In addition, we demonstrate a potential role of TGF-beta as an important modifier of receptor activator of NF-kappaB ligand (RANKL)-dependent osteoclast activation and osteolysis. Together, these studies demonstrate that inhibition of TGF-betaRI signaling at the TB interface will be a therapeutic target in the treatment of breast cancer-induced osteolysis.

Topics: Animals; Bone Neoplasms; Cell Differentiation; Cell Line, Tumor; Female; Mammary Neoplasms, Experimental; Mice; Mice, Inbred BALB C; Osteoclasts; Osteolysis; Protein Serine-Threonine Kinases; RANK Ligand; Receptor, Transforming Growth Factor-beta Type I; Receptors, Transforming Growth Factor beta; Signal Transduction; Transforming Growth Factor beta

Results recently reported in the literature have raised some concerns regarding the use of recombinant human bone morphogenetic protein (rhBMP-2) in the cervical spine.. We undertook a radiological and clinical review of cervical fusions performed at our institution with polyetheretherketone (PEEK) interbody cage and rhBMP-2.. Observational study.. Perioperative clinical and radiologic data of all patients who underwent an anterior cervical discectomy and fusion using PEEK and rhBMP-2 for cervical spondylotic radiculopathy or myelopathy were collected.. Images were examined for fusion, heterotopic ossification, end-plate resorption, subsidence, and segmental sagittal alignment.. All patients underwent detailed postoperative radiologic analysis using a computed tomography (CT) scan obtained at least 6 months postoperatively and plain X-rays obtained at regular intervals.. Twenty-two patients had 38 levels fused using PEEK and varying doses of rhBMP-2. No anterior cervical swelling requiring additional procedures or longer than anticipated hospital stays occurred. Pseudoarthrosis, shown as a horizontal radiolucent fissure through the midportion of the PEEK cage on CT, occurred in four patients. Excessive bone growth into the spinal canal or foramina occurred in 26 (68%) patients but did not result in neurologic sequelae. Cystic regions in the core of the PEEK spacer were seen in most patients, with 15 levels (39%) having cysts measuring 3mm or greater. Moderate or severe osteolysis of the end plates occurred in 57% of levels, and this led to subsidence of the construct and loss of some of the segmental sagittal alignment (ie, lordosis) that had been achieved with surgery.. The unlimited supply of PEEK spacers and rhBMP-2 and their ease of use make them attractive platforms to achieve fusion. This study has demonstrated that the fusion process using rhBMP-2 is a dynamic one, with osteolysis dominating the initial phase, leading to end-plate resorption and consequently loss of some of the disc space height and sagittal alignment that was achieved with surgery. There is a high incidence of bone growth beyond the core of the PEEK spacer and cystic regions within the cage. Given our experience, we currently reserve the use of PEEK and rhBMP-2 for use in those patients who are at greatest risk of pseudoarthrosis.

Topics: Adult; Aged; Benzophenones; Bone Cysts; Bone Morphogenetic Protein 2; Bone Morphogenetic Proteins; Bone Plates; Cervical Vertebrae; Female; Humans; Ketones; Male; Middle Aged; Osseointegration; Ossification, Heterotopic; Osteolysis; Polyethylene Glycols; Polymers; Postoperative Complications; Prosthesis Failure; Pseudarthrosis; Radiography; Recombinant Proteins; Spinal Fusion; Transforming Growth Factor beta

Increased transforming growth factor-beta (TGF-beta) signaling has been observed at the tumor-bone interface of mammary tumor-induced osteolytic lesions despite no observed transcriptional up-regulation of TGF-beta. To this point, the mechanism for enhanced TGF-beta signaling remains unclear. The bulk of TGF-beta that is released at the tumor-bone interface is in an inactive form secondary to association with beta-latency-associated protein and latency TGF-beta binding protein. We hypothesized that the observed increase in TGF-beta signaling is due to increased cathepsin G-dependent, matrix metalloproteinase 9 (MMP9)-mediated activation of latent TGF-beta. MMP9 is capable of activating latent TGF-beta, and we observed that decreased production of MMP9 was associated with reduced TGF-beta signaling. Similar to TGF-beta, MMP9 is released in an inactive form and requires proteolytic activation. We showed that cathepsin G, which we have previously shown to be up-regulated at the tumor-bone interface, is capable of activating pro-MMP9. Inhibition of cathepsin G in vivo significantly reduced MMP9 activity, increased the ratio of latent TGF-beta to active TGF-beta, and reduced the level of TGF-beta signaling. Our proposed model based on these results is that cathepsin G is up-regulated through tumor-stromal interactions and activates pro-MMP9, active MMP9 cleaves and releases active TGF-beta, and active TGF-beta can then promote tumor growth and enhance osteoclast activation and subsequent bone resorption. Thus, for the first time, we have identified cathepsin G and MMP9 as proteases involved in enhanced TGF-beta signaling at the tumor-bone interface of mammary tumor-induced osteolytic lesions and have identified these proteases as potential therapeutic targets.

Topics: Animals; Bone and Bones; Bone Neoplasms; Cathepsin G; Enzyme Activation; Enzyme Precursors; Mammary Tumor Virus, Mouse; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Mice; Oligonucleotides, Antisense; Osteolysis; RANK Ligand; Signal Transduction; Transforming Growth Factor beta

Metastatic prostate tumors in the bone microenvironment stimulate bone resorption, resulting in release of growth factors from the bone matrix that play important roles in tumor growth and osteoclast induction. Transforming growth factor beta (TGFbeta) is one of the most abundantly stored cytokines in bone matrix, regulating diverse biological activities. Here we evaluate its involvement in prostate tumor growth in the bone microenvironment, comparing with tumor growth in the subcutaneous microenvironment as a control. Rat prostate tumors were transplanted onto the cranial bone and into the subcutis of F344 male rats. Tumor cell proliferation, apoptosis, and TGFbeta signal transduction were compared between the tumor-bone interface and the tumor-subcutaneous interface. Effects of TGFbeta on osteoclast differentiation were also evaluated in vitro. Inhibitory effects of TGFbeta receptor 1 antisense oligonucleotide on TGFbeta signaling, osteolysis, osteoblasts, and tumor growth were examined in vivo. Osteolytic changes were extensively observed at the tumor-bone interface, where the TGFbeta level, TGFbeta signal transduction, and tumor cell proliferation were higher than at the tumor-subcutaneous interface. In vitro treatment with receptor activator of nuclear factor-kappaB ligand induced osteoclast differentiation of bone marrow stromal cells, and additional exposure to TGFbeta exerted promotive effects on osteoclast induction. Intratumoral injection of TGFbeta receptor 1 antisense oligonucleotide significantly reduced TGFbeta signal transduction, osteolysis, induction of osteoclast and osteoblast, and tumor cell proliferation. Thus, we experimentally show that TGFbeta derived from bone matrix promotes cell proliferation of rat prostate cancer and osteoclast activation-associated osteolysis in the bone microenvironment.

Topics: Animals; Bone and Bones; Bone Matrix; Cell Differentiation; Cell Proliferation; Cells, Cultured; Immunohistochemistry; Male; Osteoclasts; Osteolysis; Prostatic Neoplasms; Rats; Receptors, Transforming Growth Factor beta; Signal Transduction; Transforming Growth Factor beta

Bone metastases develop in approximately 30% of patients with RCC, and the mechanisms responsible for this phenomenon are unknown. We found that TGF-beta1 stimulation of RCC bone metastasis cells promotes tumor growth and bone destruction possibly by stimulating paracrine interactions between tumor cells and the bone.. Bone metastasis is a frequent complication and causes marked morbidity in patients with renal cell carcinoma (RCC). Surprisingly, the specific mechanisms of RCC interaction with bone have been scarcely studied despite the inability to prevent or effectively treat bone metastasis. Bone is a reservoir for various growth factors including the pleiotropic cytokine TGF-beta1. TGF-beta1 has been shown to have tumor-supportive effects on advanced cancers and evidence suggests its involvement in promoting the development of breast cancer bone metastasis. Here, we studied the potential role of TGF-beta1 in the growth of RCC bone metastasis (RBM).. To inhibit TGF-beta1 signaling, RBM cells stably expressing a dominant-negative (DN) TGF-betaRII cDNA were generated. The in vivo effect of TGF-beta1 on RBM tumor growth and osteolysis was determined by histological and radiographic analysis, respectively, of athymic nude mice after intratibial injection of parental, empty vector, or DN RBM cells. The in vitro effect of TGF-beta1 on RBM cell growth was determined after TGF-beta1 treatment by MTT assay.. TGF-beta1 and the TGF-beta receptors I and II (TGF-betaRI/II) were consistently expressed in both RBM tissues and cell lines. Inhibition of TGF-beta1 signaling in RBM cells significantly reduced tumor establishment and osteolysis observed in vivo after injection into the murine tibia, although no effect on tumor establishment was observed after injection of RBM cells subcutaneously or into the renal subcapsule. Treatment of five RBM cell lines with TGF-beta1 in vitro either had no effect (2/5) or resulted in a significant inhibition (3/5) of cell growth, suggesting that TGF-beta1 may promote RBM tumor growth indirectly in vivo.. TGF-beta1 stimulation of RBM cells plays a role in promoting tumor growth and subsequent osteolysis in vivo, likely through the initiation of tumor-promoting paracrine interactions between tumor cells and the bone microenvironment. These data suggest that inhibition of TGF-beta1 signaling may be useful in the treatment of RBM.

Topics: Animals; Bone Neoplasms; Carcinoma, Renal Cell; Cell Division; Cell Line, Tumor; DNA Primers; Enzyme-Linked Immunosorbent Assay; Humans; Kidney Neoplasms; Male; Mice; Mice, Nude; Osteolysis; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; Recombinant Proteins; Reverse Transcriptase Polymerase Chain Reaction; Transfection; Transforming Growth Factor beta

Although a majority of metastatic prostate cancer lesions are osteoblastic in nature, some are mixed or lytic; and, osteoblastic lesions require osteolytic activity in order to progress. The role of BMPs in the formation of prostate cancer metastases to bone remains unknown. We hypothesized that BMPs influence the development and progression of osteolytic prostate cancer lesions.. RT-PCR and Western blot analysis were used to determine BMP receptor expression on the osteolytic prostate cancer cell line PC-3. Migration, invasion, and cellular proliferation assays were performed on PC-3 cells to quantify the effects of BMP-2, -4, and -7. In vivo, PC-3 cells were injected alone, with an empty retroviral vector, or with a retroviral vector overexpressing noggin, into the tibias of SCID mice. The animals were followed for 8 weeks, and histologic and radiographic analysis were performed at 2, 4, 6, and 8 weeks.. BMP receptors are expressed on PC-3 cells, suggesting that they would be responsive to host BMP secretion. BMP-2, and to a lesser extent, BMP-4, stimulated PC-3 cell migration and invasion in a dose-dependent fashion. Noggin inhibited cellular migration and invasion of BMP-2 and -4 stimulated PC-3 cells. BMP-2 alone stimulated PC-3 cell proliferation, but BMP-4 had no effect. BMP-7 had no effect on proliferation, migration, or invasion. PC-3 cells implanted into SCID mouse tibias formed osteolytic lesions as early as 2 weeks and completely destroyed the proximal tibia by 8 weeks. Overexpression of noggin in PC-3 cells inhibited the expansion of the lesion in vivo.. BMPs influence the formation of the osteolytic prostate cancer metastases, and treatment modalities that inhibit BMP activity may limit the progression of the lytic component of prostate cancer metastases.

Topics: Animals; Bone Morphogenetic Protein 2; Bone Morphogenetic Protein 4; Bone Morphogenetic Protein 7; Bone Morphogenetic Protein Receptors; Bone Morphogenetic Proteins; Bone Neoplasms; Carrier Proteins; Cell Line, Tumor; Cell Movement; Cell Proliferation; Disease Progression; Gene Expression; Humans; Male; Mice; Mice, SCID; Neoplasm Metastasis; Neoplasm Transplantation; Osteoclasts; Osteolysis; Prostatic Neoplasms; Rats; Transfection; Transforming Growth Factor beta; Xenograft Model Antitumor Assays

Treatment of segmental bone loss remains a challenge in skeletal repairs. This study was performed to evaluate the efficacy of the use of recombinant bone morphogenetic protein-2 (rhBMP-2) delivered in an injectable calcium phosphate cement (alpha bone substitute material [alpha-BSM]) to bridge critical-sized defects in the rabbit radius.. Unilateral 20-mm mid-diaphyseal defects were created in the radii of thirty-six skeletally mature New Zealand White rabbits. The defects in twelve rabbits each were filled with 0.166 mg/mL rhBMP-2/alpha-BSM cement, 0.033 mg/mL rhBMP-2/alpha-BSM cement, or buffer/alpha-BSM cement. Six rabbits from each group were killed at four weeks, and six were killed at eight weeks. Serial radiographs were made to monitor defect-bridging and residual alpha-BSM carrier. A semiquantitative histological scoring system was used to evaluate defect-bridging. Histomorphometry was used to quantify residual alpha-BSM; trabecular bone area; trabecular bone volume fraction; and cortical length, width, and area.. At four weeks, there had been more rapid resorption of alpha-BSM and filling of the defects with trabecular bone in the group treated with 0.166 mg/mL rhBMP-2/alpha-BSM than in the other two groups. Histomorphometry confirmed an increased trabecular area and volume fraction in this group compared with the other two groups. In both rhBMP-2/alpha-BSM-treated groups, the majority of the trabecular bone was formed by a direct process adjacent to the resorbing alpha-BSM. At eight weeks, complete cortical bridging and regeneration of the marrow space were present in all of the defects treated with 0.166 mg/mL rhBMP-2/alpha-BSM. That group also had reduced residual alpha-BSM and trabecular area and volume, compared with the other two groups, at eight weeks as a result of a rapid remodeling process.. Treatment of a critical-sized defect in a rabbit radius with 0.166 mg/mL rhBMP-2/alpha-BSM injectable cement can result in bridging with cortical bone and a regenerated bone-marrow space by eight weeks. Site-specific remodeling appears to be responsible for corticalization and marrow regeneration.. RhBMP-2 delivered in a calcium phosphate cement may be useful to achieve bridging of critical-sized defects in patients. Its injectable properties may allow minimally invasive use. Delayed percutaneous administration would also be possible when augmentation is desired following an initial surgical procedure or when soft-tissue injuries preclude adequate initial treatment.

Topics: Animals; Bone Cements; Bone Morphogenetic Protein 2; Bone Morphogenetic Proteins; Bone Regeneration; Bone Substitutes; Calcium Phosphates; Disease Models, Animal; Osteolysis; Rabbits; Radiography; Radius; Recombinant Proteins; Transforming Growth Factor beta

We investigated the molecular basis for osteolytic bone metastasis by selecting human breast cancer cell line subpopulations with elevated metastatic activity and functionally validating genes that are overexpressed in these cells. These genes act cooperatively to cause osteolytic metastasis, and most of them encode secreted and cell surface proteins. Two of these genes, interleukin-11 and CTGF, encode osteolytic and angiogenic factors whose expression is further increased by the prometastatic cytokine TGF beta. Overexpression of this bone metastasis gene set is superimposed on a poor-prognosis gene expression signature already present in the parental breast cancer population, suggesting that metastasis requires a set of functions beyond those underlying the emergence of the primary tumor.

Topics: Animals; Bone Neoplasms; Breast Neoplasms; Chromatin; Connective Tissue Growth Factor; Female; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Humans; Immediate-Early Proteins; Insulin-Like Growth Factor Binding Proteins; Intercellular Signaling Peptides and Proteins; Interleukin-11; Mice; Mice, Inbred BALB C; Mice, Nude; Mitogens; Neoplasm Invasiveness; Neoplasm Proteins; Nucleic Acid Hybridization; Oligonucleotide Array Sequence Analysis; Osteolysis; Precipitin Tests; Transforming Growth Factor beta; Tumor Cells, Cultured

Bone metastasis from primary tumors is a clinically important complication of neoplastic progression. The role of parathyroid hormone-related protein (PTHrP) and transforming growth factor (TGF)-beta1 in this process has been clearly established. The current study describes an in vivo model of 13762 rat mammary carcinoma tumor cell-induced osteolysis in which PTHrP and TGF-beta1 expression is observed. Exposure of in vitro-cultured 13762 cells to doxorubicin, cis-platinum, carboplatin, methotrexate, 5-fluorouracil, paclitaxel, alendronate, risedronate, or pamidronate for 72 h resulted in varying effects on cell proliferation (IC(50) values of 0.005, 0.4, 1.9, >40, 17.9, 0.003, >40, >40, and 33.6 micro M, respectively). Tumor cells were implanted into the intramedullary space of the proximal tibia of rats, and the time course of tumor progression was evaluated using radiographic and microcomputed tomography scanning techniques. Trabecular bone mineral density, cortical bone mineral density, and whole bone mineral density were measured (in mg/cm(3)). In untreated animals, radiographic evidence of osteolysis was evident 7 days after implantation. Trabecular bone mineral density and whole bone mineral density were significantly decreased by 21 days after implantation (48% and 26%, respectively). Bisphosphonates showed broad protective activity against tumor-driven osteolysis, Immunohistochemical evaluation of s.c. and intratibially implanted cells demonstrated the expression of PTHrP and TGF-beta1. The results of this study demonstrate the ability of 13762 rat mammary carcinoma cells to elicit a measurable osteolysis and that bisphosphonates inhibit the tumor-induced bone resorption in this model.

Topics: Animals; Antineoplastic Agents; Bone Resorption; Cell Division; Cell Line, Tumor; Cell Survival; Diphosphonates; Disease Progression; Female; Humans; Mammary Neoplasms, Experimental; Neoplasm Metastasis; Osteolysis; Parathyroid Hormone-Related Protein; Rats; Rats, Inbred F344; Transforming Growth Factor beta

Orthotopic or intracardiac injection of human breast cancer cell lines into immunocompromised mice allows study of the molecular basis of breast cancer metastasis. We have established a quantitative real-time PCR approach to analyze metastatic spread of human breast cancer cells inoculated into nude mice via these routes. We employed MDA-MB-231 human breast cancer cells genetically tagged with a bacterial beta-galactosidase (Lac-Z) retroviral vector, enabling their detection by TaqMan real-time PCR. PCR detection was linear, specific, more sensitive than conventional PCR, and could be used to directly quantitative metastatic burden in bone and soft organs. Attesting to the sensitivity and specificity of the PCR detection strategy, as few as several hundred metastatic MDA-MB-231 cells were detectable in 100 microns segments of paraffin-embedded lung tissue, and only in samples adjacent to sections that scored positive by histological detection. Moreover, the measured real-time PCR metastatic burden in the bone environment (mouse hind-limbs, n = 48) displayed a high correlation to the degree of osteolytic damage observed by high resolution X-ray analysis (r2 = 0.972). Such a direct linear relationship to tumor burden and bone damage substantiates the so-called 'vicious cycle' hypothesis in which metastatic tumor cells promote the release of factors from the bone which continue to stimulate the tumor cells. The technique provides a useful tool for molecular and cellular analysis of human breast cancer metastasis to bone and soft organs, can easily be extended to other cell/marker/organ systems, and should also find application in preclinical assessment of anti-metastatic modalities.

Topics: Animals; Bone Matrix; Bone Neoplasms; Bone Resorption; Breast Neoplasms; Carcinoma, Ductal, Breast; Computer Systems; Cytokines; DNA, Neoplasm; Female; Genes, Reporter; Growth Substances; Heart; Humans; Injections; Lac Operon; Lung Neoplasms; Mammary Glands, Animal; Mice; Mice, Nude; Neoplasm Proteins; Organ Specificity; Osteoclasts; Osteolysis; Paracrine Communication; Parathyroid Hormone-Related Protein; Polymerase Chain Reaction; Proteins; Radiography; Sensitivity and Specificity; Transforming Growth Factor beta; Transplantation, Heterologous; Tumor Cells, Cultured

Extracellular matrix (ECM) remodeling is critical to morphogenesis and homeostasis. The identification of inactivating mutations in a gene encoding one of its modifying enzymes, matrix metalloproteinase 2 (MMP-2), in people with a hereditary disorder in which the bones disintegrate, represents the first genetic evidence that the proteolysis of the ECM mediates human growth and development. It also underscores the need for an intricate balance between breakdown and deposition of the ECM.

Topics: Abnormalities, Multiple; Arthritis; Extracellular Matrix; Humans; Matrix Metalloproteinase 2; Osteolysis; Syndrome; Transforming Growth Factor beta

To assess the expression of potential osteoclastogenic and osteolytic factors in osteolytic lesions from patients with Langerhans cell histiocytosis.. Paraffin-embedded biopsy sections from 5 such archival cases underwent immunohistochemical procedures with antibodies to detect the following antigens: CD(1a), S100 protein, interleukin 11, the latency-associated peptide of transforming growth factor beta(1), and angiotensin-converting enzyme.. Commonalities noted include (1) the presence of multinucleated osteoclast-like giant cells, (2) the expression of interleukin 11 and latency-associated peptide antigens in lesional Langerhans cells, and (3) plasmalemmal immunoreactivity for angiotensin-converting enzyme antigen on non-Langerhans cell histiocytes and, on occasion, osteoclast-like giant cells and endothelial cells.. These observations suggest a possible pathogenetic sequence for osteolysis in Langerhans cell histiocytosis that involves angiotensin II formation, leading to the activation of latent transforming growth factor beta(1) and, in turn, to the enhanced production of interleukin 11, resulting in both osteoclastogenesis and impaired remodeling of bone.

Topics: Adolescent; Adult; Antigens, CD1; Biopsy; Child; Endothelium; Female; Histiocytes; Histiocytosis, Langerhans-Cell; Humans; Interleukin-11; Langerhans Cells; Male; Osteoclasts; Osteolysis; Peptidyl-Dipeptidase A; S100 Proteins; Tissue Embedding; Transforming Growth Factor beta

Multiple myeloma is associated with unbalanced bone remodeling causing lytic bone lesions. Interleukin-11 (IL-11) promotes osteoclast formation and inhibits osteoblast activity and may, thus, be one factor involved in cancer-induced bone destruction. We have previously shown that myeloma cells produce hepatocyte growth factor (HGF). We now report that HGF induces IL-11 secretion from human osteoblast-like cells and from the osteosarcoma cell lines Saos-2 and HOS. In coculture experiments, both the myeloma cell line JJN-3 and primary myeloma cells from 3 patients induced IL-11 secretion from osteoblasts, whereas no induction was observed with the non-HGF producing myeloma cell line OH-2. Enhanced IL-11 induction was observed with physical contact between osteoblasts and myeloma cells as compared with experiments in which contact was prohibited by tissue inserts. Anti-HGF serum strongly reduced the myeloma cell-induced IL-11 secretion. Furthermore, we show that JJN-3 cells express HGF on the cell-surface. Removal of surface-bound HGF on JJN-3 cells reduced IL-11 production induced in cocultures. Transforming growth factor beta1 and IL-1 potentiated the effect of HGF on IL-11 secretion, whereas an additive effect was observed with tumor necrosis factor. Thus, myeloma-derived HGF can influence the bone marrow environment both as a soluble and a surface-bound factor. Furthermore, HGF emerges as a possible factor involved in myeloma bone disease by its ability to induce IL-11.

Topics: Hepatocyte Growth Factor; Humans; Interleukin-1; Interleukin-11; Multiple Myeloma; Osteoblasts; Osteolysis; Transforming Growth Factor beta; Tumor Cells, Cultured

The interactions of the cells in the bone microenvironment play important roles in bone remodeling. Osteoblasts are involved in the bone remodeling through the production of soluble factors that regulate proliferation and differentiation of osteoclasts and through cell-cell interactions. Histological studies have suggested that endothelial cells are also associated with some osteolytic bone diseases. However, it is still unclear how endothelial cells contribute to bone resorption. We established bone-derived endothelial cells (BDECs) to study their roles in bone remodeling. The established BDECs promoted bone resorption in a murine neonatal calvaria organ culture system by secreting a soluble bone resorption-inducing factor(s) when stimulated by several inflammatory cytokines. This bone resorption-inducing factor was identified as interleukin-11 (IL-11). IL-11 is known to enhance bone resorption by promoting osteoclastogenesis and by suppressing the activity of osteoblasts. The production of IL-11 in BDECs was also promoted by conditioned medium of human melanoma A375M cells. Because A375M cells formed osteolytic bone metastasis in vivo, BDECs might be involved in pathological osteolysis by producing IL-11. These results suggest that endothelial cells in bone play important roles in the promotion of bone resorption by secreting IL-11 in physiological and pathological conditions.

Topics: Animals; Bone Marrow Cells; Bone Neoplasms; Cell Line, Transformed; Cells, Cultured; Culture Media, Conditioned; Endothelium; Femur; Humans; Interleukin-1; Interleukin-11; Knee Joint; Mice; Mice, Inbred BALB C; Neoplasm Metastasis; Osteolysis; Tetradecanoylphorbol Acetate; Transforming Growth Factor beta; Tumor Cells, Cultured

Levels of bone remodeling agents were measured in conditioned media from cultures of periprosthetic pseudosynovial membranes and related to the radiographic features of the failed joint implants. Radiographs of both cemented hip (n = 28) and cemented knee (n = 11) implants were examined and the pattern of radiolucency was classified as erosive linear, or mixed. Similar levels of interleukin-1-beta (IL-1 beta), interleukin-6, tumor necrosis factor alpha (TNF-alpha), transforming growth factor beta-1, and prostaglandin E2 (PGE2) were found in pseudosynovial membrane conditioned media from all 3 groups of hips and the knee group (all linear). Significant correlations were evident only between PGE2 and TNF-alpha levels in the linear hip group and PGE2 and IL-1 beta levels in the knee group. A close correlation was found between IL-1 beta and TNF-alpha in both linear and erosive hips. It is suggested that coregulation of these bone remodeling agents differs with the radiographic appearance of the failed joint implants. As all the implants were cemented and the results contrast with those of others obtained with pseudosynovial membrane conditioned media from cementless implants, it is considered that cement critically influences the process of implant failure.

Topics: Adult; Aged; Aged, 80 and over; Cementation; Culture Media, Conditioned; Dinoprostone; Female; Growth Substances; Hip Prosthesis; Humans; Interleukin-1; Interleukin-6; Knee Prosthesis; Male; Middle Aged; Osteolysis; Prosthesis Failure; Synovial Membrane; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

The pathogenesis of breast cancer-induced osteolysis remains largely unknown. To evaluate the potential role of osteoblasts as target cells during this process, we incubated SaOS-2 human osteoblast-like cells (OBL) with culture media conditioned by proliferative (PM, 'Proliferation Media') or confluent (CfM, 'Confluence Media') MCF-7 human breast cancer cells. CfM decreased the growth of OBL by 26% (P < 0.01) while PM was without significant effect on this parameter. In contrast, both PM and CfM obtained from MCF-7 cultures increased the cyclic AMP (cAMP) response of OBL to the osteolytic agents PTH (10(-8) M) and PTH-related peptide (PTHrP, 10(-8) M) by a factor of about 3 (P < 0.001), and to prostaglandin E(2) (PGE(2),10(-6) M) by a factor of about 2 (P < 0.01). No significant modulation of OBL growth or sensitivity to PTH, PTHrP, or PGE2 was induced by media obtained from HBL-100 non-malignant immortalized breast epithelial cell cultures. 17betaestradiol (E(2), 10(-8) M) and the antiestrogen tamoxifen (Tam, 10(-7) M) added for 48 h to MCF-7 cultures before collecting conditioned media attenuated and potentiated, respectively, the PM- but not the CfM-induced increase in the response of OBL to PTH or PTHrP Along the same line, the addition to MCF-7 conditioned media of a polyclonal anti-transforming growth factor-beta (TGF-beta) antibody attenuated by about 25% (P < 0.01) the PM-induced increase in OBL response to PTH and PTHrP while abrogating the modulatory effects of E(2) and Tam on that response. Together, our results indicate that MCF-7 breast cancer cells secrete factors which inhibit the growth of OBL and increase their sensitivity to various osteolytic agents. TGF-beta was only partly responsible for these effects, and accounts for their modulation by E(2) and Tam. The identification of other osteoblast-modulatory factor(s) should contribute to a better understanding and treatment of breast cancer-induced osteolysis.

Topics: Adenocarcinoma; Breast Neoplasms; Cell Division; Colforsin; Culture Media, Conditioned; Cyclic AMP; Dinoprostone; Female; Humans; Osteoblasts; Osteolysis; Parathyroid Hormone; Parathyroid Hormone-Related Protein; Proteins; Transforming Growth Factor beta; Tumor Cells, Cultured

Osteolysis has become a major cause of aseptic loosening in total joint arthroplasty (TJA). Titanium, cobalt and chromium are commonly used in orthopaedic implants (e.g. joint prostheses). The release of bone-associated cytokines has been associated with the development of osteolysis in patients with prostheses. We evaluated the effects of these metals on the release of bone-associated cytokines (IL-1 beta, IL-6, TNF-alpha and TGF-beta 1) by human blood monocytes/macrophages and monocyte-like U937 cells upon lipopolysaccharide (LPS) stimulation, the cell proliferation, and their cytotoxic effects on these cells in vitro. We found that the release of IL-1 beta was enhanced by titanium, chromium and cobalt, the release of TNF-alpha was enhanced by titanium and chromium, and the release of IL-6 was enhanced by titanium. All three metal ions inhibited the release of TGF-beta 1. We also found that titanium and chromium, but not cobalt, enhanced blood monocyte/macrophage proliferation in response to LPS while only titanium enhanced U937 cell proliferation in response to LPS. The metals in concentrations ranging from 0.01 to 100 ngml-1 did not stimulate the cells to secrete detectable cytokines in the absence of LPS. Furthermore, a 4-h pre-exposure of blood monocytes/macrophages or U937 cells to the metals did not alter cytokine release when the metals were removed from the media prior to the addition of LPS. Similarly, a 4-h pre-exposure of blood monocytes/macrophages or U937 cells to LPS did not alter cytokine release when LPS was removed from the media prior to the addition of the metals. The metals did not reduce cell viability and induce cell injury after 72h incubation with the cells. The data suggest that the three metals at clinically relevant concentrations modulated cytokine expression, whereas they did not induce any cytotoxic effects. A metal-induced enhancement of bone-resorbing cytokine release with a concomitant inhibition of bone-forming cytokine release may be an important factor in the development of osteolysis, which can severely compromise the outcome of TJA.

Topics: Biocompatible Materials; Cell Division; Cells, Cultured; Chromium; Cobalt; Corrosion; Cytokines; Humans; Interleukin-1; Interleukin-6; Joint Prosthesis; Lymphoma, Large B-Cell, Diffuse; Macrophages; Monocytes; Osteolysis; Prosthesis Failure; Titanium; Transforming Growth Factor beta; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha

The concentrations of transforming growth factor-beta (TGF-beta) in platelets, plasma and urine from 27 patients with multiple myeloma (MM) and from 22 normal controls were measured by sandwich enzyme-linked immunosorbent assay using a monoclonal antibody specific for human TGF-beta 1+2+3. A significantly increased intraplatelet TGF-beta (24.6 +/- 9.6 vs. 17.8 +/- 8.8 ng/10(5) platelets, p < 0.01) and urinary TGF-beta (1.4 +/- 0.8 vs 1.0 +/- 0.4 ng/mg Cr, p < 0.02) were observed in MM patients compared with normal controls. The mean platelet TGF-beta level in MM patients with osteolytic lesions was more increased than in those without osteolytic lesions (29.6 +/- 7.7 vs 18.4 +/- 8.2 ng/10(5) platelets, p < 0.001). In this study, we evaluated the possible role of TGF-beta in the pathogenesis of myeloma, especially in osteolytic lesions.

Topics: Aged; Aged, 80 and over; Antibodies, Monoclonal; Blood Platelets; Enzyme-Linked Immunosorbent Assay; Female; Hematocrit; Humans; Male; Middle Aged; Multiple Myeloma; Osteolysis; Sensitivity and Specificity; Transforming Growth Factor beta; Tumor Cells, Cultured