Investigating the efficacy of bisphosphonates treatment against multiple myeloma induced bone disease using a computational model

Multiple myeloma (MM)-induced bone disease is mortal for most MM patients. Bisphosphonates are first-line treatment for MM-induced bone disease, since it can inhibit osteoclast activity and the resultant bone resorption by suppressing the differentiation of osteoclast precursors into mature osteoclasts, promoting osteoclast apoptosis and disrupting osteoclast function. However, it is still unclear whether bisphosphonates have an anti-tumour effect. In our previous work, a computational model was built to simulate the pathology of MM-induced bone disease. This paper extends this proposed computational model to investigate the efficacy of bisphosphonates treatment and then clear the controversy of this therapy. The extended model is validated through the good agreement between simulation results and experimental data. The simulation results suggest that bisphosphonates indeed have an anti-tumour effect

Osteoprotegerin reduces osteoclast resorption activity without affecting osteogenesis on nanoparticulate mineralized collagen scaffolds.

The instructive capabilities of extracellular matrix-inspired materials for osteoprogenitor differentiation have sparked interest in understanding modulation of other cell types within the bone regenerative microenvironment. We previously demonstrated that nanoparticulate mineralized collagen glycosaminoglycan (MC-GAG) scaffolds efficiently induced osteoprogenitor differentiation and bone healing. In this work, we combined adenovirus-mediated delivery of osteoprotegerin (AdOPG), an endogenous anti-osteoclastogenic decoy receptor, in primary human mesenchymal stem cells (hMSCs) with MC-GAG to understand the role of osteoclast inactivation in augmentation of bone regeneration. Simultaneous differentiation of osteoprogenitors on MC-GAG and osteoclast progenitors resulted in bidirectional positive regulation. AdOPG expression did not affect osteogenic differentiation alone. In the presence of both cell types, AdOPG-transduced hMSCs on MC-GAG diminished osteoclast-mediated resorption in direct contact; however, osteoclast-mediated augmentation of osteogenic differentiation was unaffected. Thus, the combination of OPG with MC-GAG may represent a method for uncoupling osteogenic and osteoclastogenic differentiation to augment bone regeneration

Aplidin (plitidepsin) is a novel anti-myeloma agent with potent anti-resorptive activity mediated by direct effects on osteoclasts

Despite recent progress in its treatment, Multiple Myeloma (MM) remains incurable and its associated bone disease persists even after complete remission. Thus, identification of new therapeutic agents that simultaneously suppress MM growth and protect bone is an unmet need. Herein, we examined the effects of Aplidin, a novel anti-cancer marine-derived compound, on MM and bone cells. In vitro, Aplidin potently inhibited MM cell growth and induced apoptosis, effects that were enhanced by dexamethasone (Dex) and bortezomib (Btz). Aplidin modestly reduced osteocyte/osteoblast viability and decreased osteoblast mineralization, effects that were enhanced by Dex and partially prevented by Btz. Further, Aplidin markedly decreased osteoclast precursor numbers and differentiation, and reduced mature osteoclast number and resorption activity. Moreover, Aplidin reduced Dex-induced osteoclast differentiation and further decreased osteoclast number when combined with Btz. Lastly, Aplidin alone, or suboptimal doses of Aplidin combined with Dex or Btz, decreased tumor growth and bone resorption in ex vivo bone organ cultures that reproduce the 3D-organization and the cellular diversity of the MM/bone marrow niche. These results demonstrate that Aplidin has potent anti-myeloma and anti-resorptive properties, and enhances proteasome inhibitors blockade of MM growth and bone destruction

The Cord Weekly (January 12, 1968)

Rab44 is an atypical Rab GTPase that contains some additional domains such as the EF-hand and coiled-coil domains as well as Rab-GTPase domain. Although Rab44 genes have been found in mammalian genomes, no studies concerning Rab44 have been reported yet. Here, we identified Rab44 as an upregulated protein during osteoclast differentiation. Knockdown of Rab44 by small interfering RNA promotes RANKL-induced osteoclast differentiation of the murine monocytic cell line, RAW-D or of bone marrow-derived macrophages (BMMs). In contrast, overexpression of Rab44 prevents osteoclast differentiation. Rab44 was localized in the Golgi complex and lysosomes, and Rab44 overexpression caused an enlargement of early endosomes. A series of deletion mutant studies of Rab44 showed that the coiled-coil domain and lipidation sites of Rab44 is important for regulation of osteoclast differentiation. Mechanistically, Rab44 affects nuclear factor of activated T-cells c1 (NFATc1) signaling in RANKL-stimulated macrophages. Moreover, Rab44 depletion caused an elevation in intracellular Ca2+ transients upon RANKL stimulation, and particularly regulated lysosomal Ca2+ influx. Taken together, these results suggest that Rab44 negatively regulates osteoclast differentiation by modulating intracellular Ca2+ levels followed by NFATc1 activation.長崎大学学位論文 学位記番号:博(医歯薬)甲第1003号 学位授与年月日:平成29年12月6日Author: Yu Yamaguchi, Eiko Sakai, Kuniaki Okamoto, Hiroshi Kajiya, Koji Okabe, Mariko Naito, Tomoko Kadowaki, Takayuki TsukubaCitation: Cellular and Molecular Life Sciences, 75(1), pp.33-48; 201

Analysis of Nkx3.1:Cre-driven Erk5 deletion reveals a profound spinal deformity which is linked to increased osteoclast activity

Extracellular signal-regulated protein kinase 5 (ERK5) has been implicated during development and carcinogenesis. Nkx3.1-mediated Cre expression is a useful strategy to genetically manipulate the mouse prostate. While grossly normal at birth, we observed an unexpected phenotype of spinal protrusion in Nkx3.1:Cre;Erk5fl/fl (Erk5fl/fl) mice by ~6–8 weeks of age. X-ray, histological and micro CT (µCT) analyses showed that 100% of male and female Erk5fl/fl mice had a severely deformed curved thoracic spine, with an associated loss of trabecular bone volume. Although sex-specific differences were observed, histomorphometry measurements revealed that both bone resorption and bone formation parameters were increased in male Erk5fl/fl mice compared to wild type (WT) littermates. Osteopenia occurs where the rate of bone resorption exceeds that of bone formation, so we investigated the role of the osteoclast compartment. We found that treatment of RANKL-stimulated primary bone marrow-derived macrophage (BMDM) cultures with small molecule ERK5 pathway inhibitors increased osteoclast numbers. Furthermore, osteoclast numbers and expression of osteoclast marker genes were increased in parallel with reduced Erk5 expression in cultures generated from Erk5fl/fl mice compared to WT mice. Collectively, these results reveal a novel role for Erk5 during bone maturation and homeostasis in vivo

The Direct Effect of Low-Magnitude High-Frequency Mechanical Vibration on Osteoclast Formation from RAW267.4 Monocytes

Low-magnitude high-frequency (LMHF) mechanical vibration has been demonstrated to enhance bone formation possibly through inhibition of osteoclastogenesis of bone. Earlier research has demonstrated osteoclast formation from RAW264.7 monocytes was inhibited by a chewing cycle mimicking vibration through inhibition of dendritic cell-specific transmembrane protein (DC-STAMP). We hypothesize that application of LMHF mechanical vibration directly inhibits osteoclast formation from RAW264.7 monocytes in a frequency dependent manner. RAW264.7 monocytes (ATCC) were cultured in alpha minimal essential medium (MEM) with 10% fetal bovine serum (FBS ) and 1% Pen/Strep at 37°C and 5% CO2. The cells were seeded at a density of 2000 cells/well in 96-well cell culture plates. After allowing growth overnight, the cells were treated with 20ng/ml receptor activator nuclear factor kappa-B ligand (RANKL) and refreshed every 2 days to induce osteoclast formation. In the meantime, the cells were subjected to a low-magnitude (0.3g acceleration) mechanical vibration at various frequencies (0, 30, 60 and 90 Hz) respectively. For each frequency group, the vibration was applied for 1 hour per day for 5 consecutive days. By the end of the 5th day, the cells were rinsed with 1X PBS and fixed in 4% formaldehyde for 5 minutes. Tartrate-resistant Acidic Phosphatase (TRAP, a marker enzyme of osteoclast) staining was performed. The TRAP+ multi nuclei (\u3e = 3) cells were counted and calculated. For statistical analysis, one-way ANOVA was used to test differences among the different frequency groups with Tukey post hoc comparison was used to compare between the groups, with p value being set at 0.05. Three days after RANKL stimulation, osteoclasts started to form from RAW264.7 monocytes, with a peak observed on the 5th day. After 5 days, the cells underwent apoptosis and death. Compared to the control group (0 Hz), the 30 Hz but not 60 Hz and 90 Hz frequencies of vibration group showed significant reduction of osteoclast formation by approximately 21% (p \u3c 0.05, n = 6). No significant difference was found among the three frequency groups. Low-magnitude high-frequency mechanical vibration directly inhibits osteoclast formation from RAW264.7 monocytes, and is frequency dependent

Osteoclast differentiation from human blood precursors on biomimetic calcium-phosphate substrates

The design of synthetic bone grafts to foster bone formation is a challenge in regenerative medicine. Understanding the interaction of bone substitutes with osteoclasts is essential, since osteoclasts not only drive a timely resorption of the biomaterial, but also trigger osteoblast activity. In this study, the adhesion and differentiation of human blood-derived osteoclast precursors (OCP) on two different micro-nanostructured biomimetic hydroxyapatite materials consisting in coarse (HA-C) and fine HA (HA-F) crystals, in comparison with sintered stoichiometric HA (sin-HA, reference material), were investigated. Osteoclasts were induced to differentiate by RANKL-containing supernatant using cell/substrate direct and indirect contact systems, and calcium (Ca++) and phosphorus (P5+) in culture medium were measured. We observed that OCP adhered to the experimental surfaces, and that osteoclast-like cells formed at a rate influenced by the micro- and nano-structure of HA, which also modulate extracellular Ca++. Qualitative differences were found between OCP on biomimetic HA-C and HA-F and their counterparts on plastic and sin-HA. On HA-C and HA-F cells shared typical features of mature osteoclasts, i.e. podosomes, multinuclearity, tartrate acid phosphatase (TRAP)-positive staining, and TRAP5b-enzyme release. However, cells were less in number compared to those on plastic or on sin-HA, and they did not express some specific osteoclast markers. In conclusion, blood-derived OCP are able to attach to biomimetic and sintered HA substrates, but their subsequent fusion and resorptive activity are hampered by surface micro-nano-structure. Indirect cultures suggest that fusion of OCP is sensitive to topography and to extracellular calcium.Preprin