Bone Organ Research Articles

Stimulation of fibroblast growth factor 23 by metabolic acidosis requires osteoblastic intracellular calcium signaling and prostaglandin synthesis.

Serum fibroblast growth factor 23 (FGF23) increases progressively in chronic kidney disease (CKD) and is associated with increased mortality. FGF23 is synthesized in osteoblasts and osteocytes; however, the factors regulating its production are not clear. Patients with CKD have decreased renal acid excretion leading to metabolic acidosis (MET). During MET, acid is buffered by bone with release of mineral calcium (Ca) and phosphate (P). MET increases intracellular Ca signaling and cyclooxygenase 2 (COX2)-induced prostaglandin production in the osteoblast, leading to decreased bone formation and increased bone resorption. We found that MET directly stimulates FGF23 in mouse bone organ cultures and primary osteoblasts. We hypothesized that MET increases FGF23 through similar pathways that lead to bone resorption. Neonatal mouse calvariae were incubated in neutral (NTL, pH = 7.44, Pco2 = 38 mmHg, [HCO3-] = 27 mM) or acid (MET, pH = 7.18, Pco2 = 37 mmHg, [HCO3-] = 13 mM) medium without or with 2-APB (50 μM), an inhibitor of intracellular Ca signaling or NS-398 (1 μM), an inhibitor of COX2. Each agent significantly inhibited MET stimulation of medium FGF23 protein and calvarial FGF23 RNA as well as bone resorption at 48 h. To exclude the potential contribution of MET-induced bone P release, we utilized primary calvarial osteoblasts. In these cells each agent inhibited MET stimulation of FGF23 RNA expression at 6 h. Thus stimulation of FGF23 by MET in mouse osteoblasts utilizes the same initial signaling pathways as MET-induced bone resorption. Therapeutic interventions directed toward correction of MET, especially in CKD, have the potential to not only prevent bone resorption but also lower FGF23 and perhaps decrease mortality.

Imaging characteristics associated with clinical outcomes in posterior reversible encephalopathy syndrome.

Posterior reversible encephalopathy syndrome (PRES) is a disorder of cerebrovascular autoregulation that can result in brain edema, hemorrhage, and infarction. We sought to investigate whether certain imaging characteristics in PRES are associated with clinically significant patient outcomes. We retrospectively reviewed all cases of PRES occurring between 2008 and 2014 at two major academic medical centers. Demographic, clinical, and radiographic data were collected. We analyzed imaging studies for vasogenic edema, hemorrhage, and diffusion restriction. We performed univariate analysis and stepwise logistic regression to assess the association between our radiologic findings of interest and clinical outcomes as defined by hospital discharge disposition and modified Rankin scale (mRS) at time of discharge. We identified 99 cases of PRES in 96 patients. The median age was 55years (IQR 30-65) and 74% were women. In 99 cases, 60% of patients had active cancer, 19% had history of bone marrow or organ transplantation, 14% had autoimmune disease, and 8% were peripartum. Imaging at clinical presentation showed extensive vasogenic edema in 39%, hemorrhage in 36%, hemorrhage with mass effect in 7%, and restricted diffusion in 16%. In our final logistic regression models, the presence of extensive vasogenic edema, hemorrhage with mass effect, or diffusion restriction was associated with worse clinical outcome as defined by both discharge disposition (OR=4.3; 95% CI: 1.4-36.3; p=0.047) and mRS (OR=3.6; 95% CI: 1.2-10.7; p=0.019). Extensive vasogenic edema, hemorrhage, and restricted diffusion on initial imaging in PRES are associated with worse clinical outcomes.

Engineering a humanized bone organ model in mice to study bone metastases.

Current in vivo models for investigating human primary bone tumors and cancer metastasis to the bone rely on the injection of human cancer cells into the mouse skeleton. This approach does not mimic species-specific mechanisms occurring in human diseases and may preclude successful clinical translation. We have developed a protocol to engineer humanized bone within immunodeficient hosts, which can be adapted to study the interactions between human cancer cells and a humanized bone microenvironment in vivo. A researcher trained in the principles of tissue engineering will be able to execute the protocol and yield study results within 4-6 months. Additive biomanufactured scaffolds seeded and cultured with human bone-forming cells are implanted ectopically in combination with osteogenic factors into mice to generate a physiological bone 'organ', which is partially humanized. The model comprises human bone cells and secreted extracellular matrix (ECM); however, other components of the engineered tissue, such as the vasculature, are of murine origin. The model can be further humanized through the engraftment of human hematopoietic stem cells (HSCs) that can lead to human hematopoiesis within the murine host. The humanized organ bone model has been well characterized and validated and allows dissection of some of the mechanisms of the bone metastatic processes in prostate and breast cancer.

Extracellular Matrix Scaffolds for Tissue Engineering and Regenerative Medicine.

The extracellular matrix is produced by the resident cells in tissues and organs, and secreted into the surrounding medium to provide biophysical and biochemical support to the surrounding cells due to its content of diverse bioactive molecules. Recently, the extracellular matrix has been used as a promising approach for tissue engineering. Emerging studies demonstrate that extracellular matrix scaffolds are able to create a favorable regenerative microenvironment, promote tissue-specific remodeling, and act as an inductive template for the repair and functional reconstruction of skin, bone, nerve, heart, lung, liver, kidney, small intestine, and other organs. In the current review, we will provide a critical overview of the structure and function of various types of extracellular matrix, the construction of three-dimensional extracellular matrix scaffolds, and their tissue engineering applications, with a focus on translation of these novel tissue engineered products to the clinic. We will also present an outlook on future perspectives of the extracellular matrix in tissue engineering and regenerative medicine.

Abstract P4-22-09: A phase 2 open label study of everolimus in combination with endocrine therapy in resistant hormone receptor-positive HER2-negative advanced breast cancer

Abstract Background: Therapies targeting estrogen receptor (ER) signaling are standard for patients (pts) with hormone receptor positive (HR+) (ER and/or progesterone receptor [PR] positive) metastatic breast cancer (MBC). Dysregulation of the mammalian target of rapamycin (mTOR) pathway has been associated with endocrine therapy (ET) resistance. BOLERO-2 demonstrated that the addition of the mTOR inhibitor, everolimus (EVE), to exemestane doubled the PFS in HR+ HER2- MBC pts who previously progressed on nonsteroidal aromatase inhibitor therapy.The premise of this phase 2 trial in HR+ MBC is that the addition of EVE to the last ET on which the disease progressed may restore sensitivity to ET and extend the benefit of the anti-estrogen therapy. Methods: Pts ≥18 yrs with HR+, HER2- unresectable, locally recurrent, or MBC refractory to ET in either the adjuvant or advanced/metastatic setting. 0-1 chemotherapy (chemo) regimens for MBC were permitted. Post-/ pre-/peri-menopausal women were eligible with ovarian function suppression permitted. Additional eligibility requirements include: no prior mTOR inhibitor therapy, measurable or evaluable disease, ECOG ≤2, adequate bone marrow and organ function. EVE (10 mg PO daily) was administered on a 4-wk cycle in combination with the same dose and schedule of the last ET to which their disease became resistant. Disease assessments were performed every 2 cycles and treatment continued until disease progression or unacceptable toxicity. Blood samples and archival tumor were collected respectively for the VeriStrat Assay and for the Foundation One molecular profiling platform. Results: 48 pts were enrolled; data from 26 pts is presented. Median age 63.5 yrs (range, 36-81) with 46% ≥ 65 yrs. 14 (54%) pts had received chemo in the adjuvant setting, 9 pts (35%) received chemo for MBC, and 4 pts (15%) received chemo in both settings. All pts had at least 1 prior hormonal therapy; 9 pts received ≥ 3 hormonal agents. EVE was combined with tamoxifen (27%), AIs (61%), and fulvestrant (12%). Median time on treatment was 18.6 wks (range 1-48.9 weeks). 5 pts (19%) remain on treatment and 21 (81%) have discontinued therapy due to: disease progression - 17, toxicity -2, and other causes - 2. 23 pts were evaluable for response. 1 pt on fulvestrant plus EVE had a PR and 18 pts (78%) had SD as best response, with SD > 6 mos in 7 pts, for a clinical benefit rate (CR+PR+ SD ≥ 6 months) of 35%. With a median follow up of 11 mos (range 2-16 mos), the median PFS was 6.6 months (range 3.6-9.4); the median OS has not been reached. Treatment-related adverse events consisted mostly of stomatitis, rash and fatigue with few G3 events: stomatitis 3 pts, rash 2 pts, and 1 each of fatigue, edema, and neutropenia. G1 pneumonitis was present in 2 pts. There were no G4 events or treatment related deaths. Conclusions: In HR+ HER2- advanced/MBC patients who progressed on prior ET, the addition of EVE to the ET to which their disease became resistant, resulted in 1 PR and 7 pts with SD > 6 mos. The results of the full study population will be presented. Modulation of the mTOR/AKT/PI3K pathway with EVE may extend the benefit of ET, even after tumor progression on ET alone. Citation Format: Yardley DA, Blakely L, Hemphill B, Joseph M, Liggett W, Daniel B, Castrellon A, Shastry M, Finney L, DeBusk L, Hainsworth JD, Burris III HA. A phase 2 open label study of everolimus in combination with endocrine therapy in resistant hormone receptor-positive HER2-negative advanced breast cancer [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P4-22-09.

Pigment Epithelium-Derived Factor (PEDF) mediates cartilage matrix loss in an age-dependent manner under inflammatory conditions

BackgroundInflammation is a major cause of cartilage destruction and leads to the imbalance of metabolic activities in the arthritic joint. Pigment epithelium-derived factor (PEDF) has been reported to have both pro- and anti-inflammatory activities in various cell types and to be upregulated in the arthritic joint, but its role in joint destruction is unclear. Our aim was to investigate the role of PEDF in cartilage degeneration under inflammatory conditions.MethodsPEDF was ectopically expressed in primary human articular chondrocytes, and catabolic gene expression and protein secretion in response to the pro-inflammatory cytokine interleukin 1 beta (IL-1β) were evaluated. Metatarsal bones from PEDF-deficient and wild type mice were cultured in the presence or absence of IL-1β. Cartilage matrix integrity and matrix metalloproteinases MMP-1, MMP-3, and MMP-13 were evaluated. PEDF-deficient and wild type mice were evaluated in the monosodium iodoacetate (MIA) inflammatory joint destruction animal model to determine the role of PEDF in inflammatory arthritis in vivo. Student’s t-tests and Mann–Whitney tests were employed where appropriate, for parametric and non-parametric data, respectively.ResultsWe showed that PEDF protein levels were higher in human osteoarthritis samples compared to normal samples. We demonstrated that ectopic PEDF expression in primary human articular chondrocytes exacerbated catabolic gene expression in the presence of IL-1β. In whole bone organ cultures, IL-1β induced MMP-1, MMP-3 and MMP-13 protein production, and caused significant cartilage matrix loss. Interestingly, Toluidine Blue staining showed that PEDF-deficient bones from 29 week old animals, but not 10 week old animals, had reduced matrix loss in response to IL-1β compared to their wild type counterparts. In addition, PEDF-deficiency in 29 week old animals preserved matrix integrity and protected against cell loss in the MIA joint destruction model in vivo.ConclusionWe conclude that PEDF exacerbates cartilage degeneration in an age-dependent manner under an inflammatory setting. This is the first study identifying a specific role for PEDF in joint inflammation and highlights the multi-faceted activities of PEDF.

Relationship between the uptake of 18F-borono-L-phenylalanine and L-[methyl-11C] methionine in head and neck tumors and normal organs

Background and purposeThe purpose of this study was to determine the distribution of 4-borono-2-18F-fluoro-phenylalanine (18F-BPA) and L-[methyl-11C] methionine (11C-Met) in normal organs and tumors and to evaluate the usefulness of 11C-Met/PET in screening potential candidates for boron neutron capture therapy (BNCT).Material methodsSeven patients who had at least one histologically confirmed head and neck tumor were included in this study. They underwent both whole-body 18F-BPA-PET/CT and 11C-Met-PET/CT within a span of 6 months. Uptake was evaluated using the maximum standardized uptake value (SUVmax). Regions of interest (ROIs) were placed within the tumors and target organs of brain, thyroid, submandibular gland, lung, liver, esophagus, stomach pancreas, spleen, muscle, and bone marrow.ResultsThe tumor SUVmax of FBPA and 11C-Met showed strong correlation (r2 = 0.72, P = 0.015). Although 18F-BPA and 11C-Met showed markedly different uptake in some organs (submandibular gland, liver, heart, stomach pancreas, spleen, and bone marrow), the uptake of 11C-Met was consistently higher than that of 18F-BPA in these cases.Conclusion11C-Met PET/CT might be used instead of 18F-BPA PET/CT to predict the accumulation of 10B in tumors and to select candidates for BNCT. However, it would not be suitable for evaluating accumulation in some normal organs. Therefore, the 18F-BPA-PET study remains a prerequisite for BNCT. This is the first report of the correlation between 18F-BPA and 11C-Met accumulation.

Glucocorticoids Induce Bone and Muscle Atrophy by Tissue-Specific Mechanisms Upstream of E3 Ubiquitin Ligases.

Glucocorticoid excess, either endogenous with diseases of the adrenal gland, stress, or aging or when administered for immunosuppression, induces bone and muscle loss, leading to osteopenia and sarcopenia. Muscle weakness increases the propensity for falling, which, combined with the lower bone mass, increases the fracture risk. The mechanisms underlying glucocorticoid-induced bone and muscle atrophy are not completely understood. We have demonstrated that the loss of bone and muscle mass, decreased bone formation, and reduced muscle strength, hallmarks of glucocorticoid excess, are accompanied by upregulation in both tissues in vivo of the atrophy-related genes atrogin1, MuRF1, and MUSA1. These are E3 ubiquitin ligases traditionally considered muscle-specific. Glucocorticoids also upregulated atrophy genes in cultured osteoblastic/osteocytic cells, in ex vivo bone organ cultures, and in muscle organ cultures and C2C12 myoblasts/myotubes. Furthermore, glucocorticoids markedly increased the expression of components of the Notch signaling pathway in muscle in vivo, ex vivo, and in vitro. In contrast, glucocorticoids did not increase Notch signaling in bone or bone cells. Moreover, the increased expression of atrophy-related genes in muscle, but not in bone, and the decreased myotube diameter induced by glucocorticoids were prevented by inhibiting Notch signaling. Thus, glucocorticoids activate different mechanisms in bone and muscle that upregulate atrophy-related genes. However, the role of these genes in the effects of glucocorticoids in bone is unknown. Nevertheless, these findings advance our knowledge of the mechanism of action of glucocorticoids in the musculoskeletal system and provide the basis for novel therapies to prevent glucocorticoid-induced atrophy of bone and muscle.

Intracranial Tumorous Lesion as the First Manifestation of Langerhans Cell Histiocytosis: A Diagnostic and Therapeutic Challenge - Case Report

Langerhans cell histiocytosis (LHC) is a rare proliferative disease of unknown etiology characterized by bone marrow-derived histiocyte-like cell proliferation. The disease is confirmed by electron microscopy or immunohistochemical reactivity of histiocytes to the CD1a receptor, langerin (CD207) and/or S100 protein. The clinical spectrum ranges from an acute disseminated disease to solitary or few chronic lesions of bone or other organs. Central nervous system invasion in LCH patients has rarely been reported, especially in the adult population. We report a case of an intracranial tumorous lesion as the first manifestation of LCH in a 23 year old Caucasian man. The patient presented a 2 year history of cognitive decline and a 7 year history of diabetes insipidus, diabetes mellitus, hypothyroidism and obesity. The histopathological findings of a lesion located in the hypothalamic region confirmed the diagnosis of LCH.

The Nationwide Study of Erdheim-Chester Disease in Japan

Background: Erdheim-Chester disease (ECD) is a subtype of non-Langerhans histiocytic disorder that is shown to be driven by hyperactivation of MAPK pathway (most frequently caused by BRAFV600E mutation) and characterized by generalized organ dysfunction with infiltration of CD68-positive, CD1a-negative histiocytes. However, ECD is a rare entity and therefore very little is known about epidemiology of this disorder.Methods: We underwent a postal questionnaire-based, multi-center retrospective study to clarify the clinical features of ECD patients. We first sent 3850 questionnaires to various departments including orthopedics, respiratory medicine, dermatology, hematology, and pathology to cover as many ECD patients as possible, and identified 71 ECD patients in Japan. We further collected detailed clinical information and patients' samples if available. All cases were pathologically proven and the diagnoses of ECD were self-reported by each institute. DNA was extracted from each clinical sample and Sanger sequencing or allele-specific polymerase chain reaction (PCR) for BRAFV600E mutation were underwent with specific primers.Results: Among 71 patients with ECD, detailed clinical information about 38 patients were collected. The median age was 51 years old (range: 25-76 years old) and there was a male predominance (1.9:1). Major affected lesions were the bone (84 %), central nervous system (CNS; 50 %), cardiovascular lesion (37 %), skin (37 %), retroperitoneum (37 %), endocrines (37 %), lung (24 %), and digestive organ (12 %). C-reactive protein at onset was higher than upper normal limit in 24 of 29 patients (median 23.8 mg/L). The median time from the onset to diagnosis was 17 months and median survival from initial onset was 10.9 years. In the univariate analyses, age older than 60 years old at initial onset (hazard ratio [HR], 30.2; 95% CI, 5.7-159; p < 0.001), weight loss (HR, 5.4; 95% CI, 1.4-20.8; p = 0.014), CNS involvement (HR, 25.2; 95% CI, 3.1-203; p = 0.0024), cardiovascular lesion (HR, 3.6; 95% CI, 1.2-10.8; p = 0.021) and digestive organ disease (HR, 5.6; 95% CI, 1.6-20.2; p = 0.0085) were associated with poor prognosis. Interestingly, the bone involvement was associated with better outcome (HR, 0.20; 95% CI, 0.065-0.63; p = 0.0052). Multivariate analysis revealed that older age was an independent poor prognostic factor (HR, 18.9; 95% CI, 2.12-169; p = 0.0085). In addition, patients with CNS involvement were significantly older than patients without CNS disease (median, 63.7 vs 44.0 years old; p < 0.001). We also analyzed the BRAF V600E mutation status in seven cases. Allele-specific PCR identified that three of seven patients had BRAFV600E mutation.Conclusion: Our nationwide survey revealed that older age was associated with CNS involvement and poor prognosis in ECD patients. In addition, the bone, cardiovascular, CNS and digestive organ involvement might also affect clinical outcome. It is of note that the bone lesion was associated with better survival in the univariate analysis. However, larger and prospective studies are warranted. Regional disparity such as percentage of bone lesions should be also investigated. DisclosuresOgura:Payment for lectures including service on speakers bureaus: Speakers Bureau.

Pathogeny of Fatigued Walking Condition in Pekin Ducks.

Fatigued walking condition (FWC) in ducks is an important welfare and processing issue during the loading/unloading to the processing plant that can be related to heart and bone development. An experiment was conducted to evaluate the effects of incubation conditions on duck embryo bone and heart development and their subsequent effects on ducks showing FWC at market age. Four groups of 2500 Pekin duck eggs were subjected to combinations of two incubation temperature profiles (elevated [E] and normal [N]) and two eggshell conductance profiles (G) (reduced [GR] and normal [GN]). At hatch 10 ducklings from each treatment combination were sampled for heart, liver, residual yolk, and total body weight as well as relative weights (organ percentage of whole body weight). Femur, tibia, and tarsus length and weight were also obtained, and relative asymmetry (RA) was calculated for each leg section. At 35 days of age during unloading of the truck at the slaughter plant, five hens and five drakes demonstrating normal walking and FWC were sampled. Body, heart, and ventricular weights were obtained along with femur, tibia and tarsus length, weight, and RA. Bone strength was evaluated using a three-point bending test, and tibia ash content was assessed. At hatch duckling bone characteristics and organ weights were found to be primarily affected by GR conditions, while heart development in older ducks was mainly impacted by E incubation temperatures. Tibia and relative weight at 35 days were also increased by GR and E. Fatigued ducks presented heavier tibias with more RA and cortical thickness but lower ash percentage. In conclusion, the changes in bone development during incubation and posthatch life were related to duck FWC presence during transportation to the processing plant.

One-Step Self-Assembling Nanomicelles for Pirarubicin Delivery To Overcome Multidrug Resistance in Breast Cancer.

Tumor cells can acquire multidrug resistance (MDR) as a result of drug efflux mediated by P-glycoprotein (P-gp). Here we report a targeted delivery system to carry pirarubicin (THP) to MDR breast cancer both in vitro and in vivo. PEG-derivatized vitamin E (PAMV6) amphiphiles loaded with THP were self-assembled in a single step. The PAMV6 micelles showed unimodal size distribution and high drug loading efficiency. Cytotoxicity of PAMV6/THP was higher than that of free THP on MCF-7/ADR cells but comparable to that of THP on MCF-7 cells. PAMV6/THP was able to reverse MDR more than free THP in MCF-7/ADR cells, likely reflecting the remarkably higher intracellular THP concentration in micelle-treated cells and PAMV6-mediated inhibition of P-gp activity. PAMV6/THP micelles were internalized into MCF-7/ADR cells via macropinocytosis and caveolin-mediated endocytosis, further avoiding P-gp-mediated efflux. Mechanistic studies revealed that blank PAMV6 micelles inhibited P-gp activity but did not affect P-gp expression, by significantly reducing mitochondrial membrane potential and slightly decreasing intracellular ATP levels. In a nude mouse xenograft model, PAMV6/THP led to much greater THP accumulation in tumors and much slower tumor growth than free THP. At the same time, PAMV6/THP was associated with significantly less severe bone marrow suppression and organ toxicity than free THP. Our results indicate that this PAMV6-based micelle system holds promise for combating MDR in cancer therapy.

Hedgehog signaling in bone regulates whole-body energy metabolism through a bone-adipose endocrine relay mediated by PTHrP and adiponectin.

Bone plays a role in energy metabolism, but the interplay between bone and other organs in this process is not completely understood. Here, we show that upregulated Hh signaling in bones results in increased whole-body energy expenditure, white adipose tissue (WAT) browning, hypoglycemia and skeletal muscle atrophy. We found that Hh signaling induces PTHrP secretion from bones and causes WAT browning. Injection of PTHrP-neutralizing antibody attenuates WAT browning and improves the circulating blood glucose level while high-fat diet treatment only rescues hypoglycemia. Furthermore, bone-derived PTHrP stimulates adiponectin secretion in WAT and results in systemic increase of fatty acid oxidation and glucose uptake. Mechanistically, PTHrP activates both PKA/cAMP and Akt/Foxo pathways for Ucp1 expression in WAT. PTHrP couples adiponectin actions to activate the AMPK pathway in the skeletal muscles and liver, respectively, for fatty acid oxidation. Our findings establish a new bone-adipose hormonal relay that regulates whole-body energy metabolism.

Incentives and Ethics in the Economics of Body Parts

Recent research shows that properly devised economic incentives increase the supply of blood without hampering its safety; similar effects may be expected also for other body parts such as bone marrow and organs. These positive effects alone, however, do not necessarily justify the introduction of payments for supplying body parts; these activities concern contested commodities or repugnant transactions, i.e. societies may want to prevent certain ways to regulate a transaction even if they increased supply, because of ethical concerns. When transactions concern contested commodities, therefore, societies often face trade-offs between the efficiency-enhancing effects of trades mediated by a monetary price, and the moral opposition to the provision of these payments. In this essay, I first describe and discuss the current debate on the role of moral repugnance in controversial markets, with a focus on markets for organs, tissues, blood and plasma. I then report on recent research focused on understanding the trade-offs that individuals face when forming their opinions about how a society should organize certain transactions.

Passive Entrapment of Tumor Cells Determines Metastatic Dissemination to Spinal Bone and Other Osseous Tissues.

During the metastatic process tumor cells circulate in the blood stream and are carried to various organs. In order to spread to different organs tumor cell—endothelial cell interactions are crucial for extravasation mechanisms. It remains unclear if tumor cell dissemination to the spinal bone occurs by passive entrapment of circulating tumor cells or by active cellular mechanisms mediated by cell surface molecules or secreted factors. We investigated the seeding of three different tumor cell lines (melanoma, lung and prostate carcinoma) to the microvasculature of different organs. Their dissemination was compared to biologically passive microbeads. The spine and other organs were resected three hours after intraarterial injection of tumor cells or microbeads. Ex vivo homogenization and fluorescence analysis allowed quantification of tumor cells or microbeads in different organs. Interestingly, tumor cell distribution to the spinal bone was comparable to dissemination of microbeads independent of the tumor cell type (melanoma: 5.646% ± 7.614%, lung: 6.007% ± 1.785%, prostate: 3.469% ± 0.602%, 7 μm beads: 9.884% ± 7.379%, 16 μm beads: 7.23% ± 1.488%). Tumor cell seeding differed significantly between tumor cells and microbeads in all soft tissue organs. Moreover, there were significant differences between the different tumor cell lines in their dissemination behaviour to soft tissue organs only. These findings demonstrate that metastatic dissemination of tumor cells to spinal bone and other osseous organs is mediated by passive entrapment of tumor cells similar to passive plugging of microvasculature observed after intraarterial microbeads injection.

The MET/Vascular Endothelial Growth Factor Receptor (VEGFR)-targeted Tyrosine Kinase Inhibitor Also Attenuates FMS-dependent Osteoclast Differentiation and Bone Destruction Induced by Prostate Cancer

The tyrosine kinase inhibitor TAS-115 that blocks VEGF receptor and hepatocyte growth factor receptor MET signaling exhibits antitumor properties in xenografts of human gastric carcinoma. In this study, we have evaluated the efficacy of TAS-115 in preventing prostate cancer metastasis to the bone and bone destruction using the PC3 cell line. When PC3 cells were injected into proximal tibiae in nude mouse, severe trabecular and cortical bone destruction and subsequent tumor growths were detected. Oral administration of TAS-115 almost completely inhibited both PC3-induced bone loss and PC3 cell proliferation by suppressing osteoclastic bone resorption. In an ex vivo bone organ culture, PC3 cells induced osteoclastic bone resorption when co-cultured with calvarial bone, but TAS-115 effectively suppressed the PC3-induced bone destruction. We found that macrophage colony-stimulating factor-dependent macrophage differentiation and subsequent receptor activator of NF-κB ligand-induced osteoclast formation were largely suppressed by adding TAS-115. The phosphorylation of the macrophage colony-stimulating factor receptor FMS and osteoclast related kinases such as ERK and Akt were also suppressed by the presence of TAS-115. Gene expression profiling showed that FMS expression was only seen in macrophage and in the osteoclast cell lineage. Our study indicates that tyrosine kinase signaling in host pre-osteoclasts/osteoclasts is critical for bone destruction induced by tumor cells and that targeting of MET/VEGF receptor/FMS activity makes it a promising therapeutic candidate for the treatment of prostate cancer patients with bone metastasis.

3D Bioprinting: 3D Bioprinting of Developmentally Inspired Templates for Whole Bone Organ Engineering (Adv. Healthcare Mater. 18/2016)

3D Bioprinting: 3D Bioprinting of Developmentally Inspired Templates for Whole Bone Organ Engineering (Adv. Healthcare Mater. 18/2016)

Cortical bone loss caused by glucocorticoid excess requires RANKL production by osteocytes and is associated with reduced OPG expression in mice.

Glucocorticoid excess is a major cause of low bone mass and fractures. Glucocorticoid administration decreases cortical thickness and increases cortical porosity in mice, and these changes are associated with increased osteoclast number at the endocortical surface. Receptor activator of NF-κB ligand (RANKL) produced by osteocytes is required for osteoclast formation in cancellous bone as well as the increase in cortical bone resorption caused by mechanical unloading or dietary calcium deficiency. However, whether osteocyte-derived RANKL also participates in the increase in bone resorption caused by glucocorticoid excess is unknown. To address this question, we examined the effects of prednisolone on cortical bone of mice lacking RANKL production in osteocytes. Prednisolone administration increased osteoclast number at the endocortical surface, increased cortical porosity, and reduced cortical thickness in control mice, but none of these effects occurred in mice lacking RANKL in osteocytes. Prednisolone administration did not alter RANKL mRNA abundance but did reduce osteoprotegerin (OPG) mRNA abundance in osteocyte-enriched cortical bone. Similarly, dexamethasone suppressed OPG but did not increase RANKL production in cortical bone organ cultures and primary osteoblasts. These results demonstrate that RANKL produced by osteocytes is required for the cortical bone loss caused by glucocorticoid excess but suggest that the changes in endocortical resorption are driven by reduced OPG rather than elevated RANKL expression.

Metal artifact reduction (MAR) based on two-compartment physical modeling: evaluation in patients with hip implants

Artifacts from metallic implants can hinder image interpretation in computed tomography (CT). Image quality can be improved using metal artifact reduction (MAR) techniques. To evaluate the impact of a MAR algorithm on image quality of CT examinations in comparison to filtered back projection (FBP) in patients with hip prostheses. Twenty-two patients with 25 hip prostheses who underwent clinical abdominopelvic CT on a 64-row CT were included in this retrospective study. Axial images were reconstructed with FBP and five increasing MAR levels (M30-34). Objective artifact strength (OAS) (SIart-SInorm) was assessed by region of interest (ROI) measurements in position of the strongest artifact (SIart) and in an osseous structure without artifact (SInorm) (in Hounsfield units [HU]). Two independent readers evaluated subjective image quality regarding metallic hardware, delineation of bone, adjacent muscle, and pelvic organs on a 5-point scale (1, non-diagnostic; 5, excellent image quality). Artifacts in the near field, far field, and newly induced artifacts due to the MAR technique were analyzed. OAS values were: M34: 243.8 ± 155.4 HU; M33: 294.3 ± 197.8 HU; M32: 340.5 ± 210.1 HU; M31: 393.6 ± 225.2 HU; M30: 446.8 ± 224.2 HU and FBP: 528.9 ± 227.7 HU. OAS values were significantly lower for M32-34 compared to FBP (P < 0.01). For overall subjective image quality, results were: FBP, 2.0 ± 0.2; M30, 2.3 ± 0.8; M31, 2.6 ± 0.5; M32, 3.0 ± 0.6; M33, 3.5 ± 0.6; and M34, 3.8 ± 0.4 (P < 0.001 for M30-M34 vs. FBP, respectively). Increasing MAR levels resulted in new artifacts in 17% of reconstructions. The investigated MAR algorithm led to a significant reduction of artifacts from metallic hip implants. The highest MAR level provided the least severe artifacts and the best overall image quality.

Generation of a Bone Organ by Human Adipose-Derived Stromal Cells Through Endochondral Ossification.

: Recapitulation of endochondral ossification (ECO) (i.e., generation of marrow-containing ossicles through a cartilage intermediate) has relevance to develop human organotypic models for bone or hematopoietic cells and to engineer grafts for bone regeneration. Unlike bone marrow-derived stromal cells (also known as bone marrow-derived mesenchymal stromal/stem cells), adipose-derived stromal cells (ASC) have so far failed to form a bone organ by ECO. The goal of the present study was to assess whether priming human ASC to a defined stage of chondrogenesis in vitro allows their autonomous ECO upon ectopic implantation. ASC were cultured either as micromass pellets or into collagen sponges in chondrogenic medium containing transforming growth factor-β3 and bone morphogenetic protein-6 for 4 weeks (early hypertrophic templates) or for two additional weeks in medium supplemented with β-glycerophosphate, l-thyroxin, and interleukin1-β to induce hypertrophic maturation (late hypertrophic templates). Constructs were implanted in vivo and analyzed after 8 weeks. In vitro, ASC deposited cartilaginous matrix positive for glycosaminoglycans, type II collagen, and Indian hedgehog. Hypertrophic maturation induced upregulation of type X collagen, bone sialoprotein, and matrix metalloproteinase13 (MMP13). In vivo, both early and late hypertrophic templates underwent cartilage remodeling, as assessed by MMP13- and tartrate-resistant acid phosphatase-positive staining, and developed bone ossicles, including bone marrow elements, although to variable degrees of efficiency. In situ hybridization for human-specific sequences and staining with a human specific anti-CD146 antibody demonstrated the direct contribution of ASC to bone and stromal tissue formation. In conclusion, despite their debated skeletal progenitor nature, human ASC can generate bone organs through ECO when suitably primed in vitro. Recapitulation of endochondral ossification (ECO) (i.e., generation of marrow-containing ossicles through a cartilage intermediate) has relevance to develop human organotypic models for bone or hematopoietic cells and to engineer grafts for bone regeneration. This study demonstrated that expanded, human adult adipose-derived stromal cells can generate ectopic bone through ECO, as previously reported for bone marrow stromal cells. This system can be used as a model in a variety of settings for mimicking ECO during development, physiology, or pathology (e.g., to investigate the role of BMPs, their receptors, and signaling pathways). The findings have also translational relevance in the field of bone regeneration, which, despite several advances in the domains of materials and surgical techniques, still faces various limitations before being introduced in the routine clinical practice.