Browsing by Subject "Osteoclast"
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Item Blocking the ZZ domain of sequestosome1/p62 suppresses myeloma growth and osteoclast formation in vitro and induces dramatic bone formation in myeloma-bearing bones in vivo(SpringerNature, 2016-02) Teramachi, Jumpei; Silbermann, Rebecca; Yang, Peng; Zhao, Wei; Mohammad, Khalid S.; Guo, Jianxia; Anderson, Judith L.; Zhou, Dan; Feng, Rentian; Myint, Kyaw-Zeyar; Maertz, Nathan; Beumer, Jan H.; Eiseman, Julie L.; Windle, Jolene J.; Xie, Xiang-Qun; Roodman, G. David; Kurihara, Noriyoshi; Department of Medicine, IU School of MedicineWe reported that p62 (sequestosome 1) serves as a signaling hub in bone marrow stromal cells (BMSCs) for the formation of signaling complexes, including NFκB, p38MAPK and JNK, that are involved in the increased osteoclastogenesis and multiple myeloma (MM) cell growth induced by BMSCs that are key contributors to multiple myeloma bone disease (MMBD), and demonstrated that the ZZ domain of p62 (p62-ZZ) is required for BMSC enhancement of MMBD. We recently identified a novel p62-ZZ inhibitor, XRK3F2, which inhibits MM cell growth and BMSC growth enhancement of human MM cells. In the current study, we evaluate the relative specificity of XRK3F2 for p62-ZZ, characterize XRK3F2's capacity to inhibit growth of primary MM cells and human MM cell lines, and test the in vivo effects of XRK3F2 in the immunocompetent 5TGM1 MM model. We found that XRK3F2 induces dramatic cortical bone formation that is restricted to MM containing bones and blocked the effects and upregulation of tumor necrosis factor alpha (TNFα), an osteoblast (OB) differentiation inhibitor that is increased in the MM bone marrow microenvironment and utilizes signaling complexes formed on p62-ZZ, in BMSC. Interestingly, XRK3F2 had no effect on non-MM bearing bone. These results demonstrate that targeting p62 in MM models has profound effects on MMBD.Item Bone Microenvironment-Suppressed T Cells Increase Osteoclast Formation and Osteolytic Bone Metastases in Mice(Wiley, 2022) Arellano, Danna L.; Juárez, Patricia; Verdugo-Meza, Andrea; Almeida-Luna, Paloma S.; Corral-Avila, Juan A.; Drescher, Florian; Olvera, Felipe; Jiménez, Samanta; Elzey, Bennett D.; Guise, Theresa A.; Fournier, Pierrick G.J.; Medicine, School of MedicineImmunotherapies use components of the immune system, such as T cells, to fight cancer cells, and are changing cancer treatment, causing durable responses in some patients. Bone metastases are a debilitating complication in advanced breast and prostate cancer patients. Approved treatments fail to cure bone metastases or increase patient survival and it remains unclear whether immunotherapy could benefit patients. The bone microenvironment combines various immunosuppressive factors, and combined with T cell products could increase bone resorption fueling the vicious cycle of bone metastases. Using syngeneic mouse models, our study revealed that bone metastases from 4T1 breast cancer contain tumor-infiltrating lymphocyte (TILs) and their development is increased in normal mice compared to immunodeficient and T-cell depleted mice. This effect seemed caused by the TILs specifically in bone, because T-cell depletion increased 4T1 orthotopic tumors and did not affect bone metastases from RM-1 prostate cancer cells, which lack TILs. T cells increased osteoclast formation ex vivo and in vivo contributing to bone metastasis vicious cycle. This pro-osteoclastic effect is specific to unactivated T cells, because activated T cells, secreting interferon γ (IFNγ) and interleukin 4 (IL-4), actually suppressed osteoclastogenesis, which could benefit patients. However, non-activated T cells from bone metastases could not be activated in ex vivo cultures. 4T1 bone metastases were associated with an increase of functional polymorphonuclear and monocytic myeloid-derived suppressor cells (MDSCs), potent T-cell suppressors. Although effective in other models, sildenafil and zoledronic acid did not affect MDSCs in bone metastases. Seeking other therapeutic targets, we found that monocytic MDSCs are more potent suppressors than polymorphonuclear MDSCs, expressing programmed cell death receptor-1 ligand (PD-L1)+ in bone, which could trigger T-cell suppression because 70% express its receptor, programmed cell death receptor-1 (PD-1). Collectively, our findings identified a new mechanism by which suppressed T cells increase osteoclastogenesis and bone metastases. Our results also provide a rationale for using immunotherapy because T-cell activation would increase their anti-cancer and their anti-osteoclastic properties.Item CD166 modulates disease progression and osteolytic disease in multiple myeloma(2016-03-16) Xu, Linlin; Xu, LinlinMultiple myeloma (MM) is an incurable malignancy characterized by the proliferation of neoplastic plasma cells in the bone marrow (BM) and by multiple osteolytic lesions throughout the skeleton. We previously reported that CD166 is a functional molecule on normal hematopoietic stem cells (HSC) that plays a critical role in HSC homing and engraftment, suggesting that CD166 is involved in HSC trafficking and lodgment. CD166, a member of the immunoglobulin superfamily capable of mediating homophilic interactions, has been shown to enhance metastasis and invasion in several tumors. However, whether CD166 is involved in MM and plays a role in MM progression has not been addressed. We demonstrated that a fraction of all human MM cell lines tested and MM patients’ BM CD138+ cells express CD166. Additionally, CD166+ cells preferentially home to the BM of NSG mice. Knocking-down (KD) CD166 expression on MM cells with shRNA reduced their homing to the BM. Furthermore, in a long-term xenograft model, NSG mice inoculated with CD166KD cells showed delayed disease progression and prolonged survival compared to mice receiving mock transduced cells. To examine the potential role of CD166 in osteolytic lesions, we first used a novel Ex Vivo Organ Culture Assay (EVOCA) which creates an in vitro 3D system for the interaction of MM cells with the bone microenvironment. EVOCA data from MM cells lines as well as from primary MM patients’ CD138+ BM cells demonstrated that bone osteolytic resorption was significantly reduced when CD166 was absent on MM cells or calvarial cells. We then confirmed our ex vivo findings with intra-tibial inoculation of MM cells in vivo. Mice inoculated with CD166KD cells had significantly less osteolytic lesions. Further analysis demonstrated that CD166 expression on MM cells alters bone remodeling by inhibiting RUNX2 gene expression in osteoblast precursors and increasing RANKL to OPG ratio in osteoclast precursors. We also identified that CD166 is indispensable for osteoclastogenesis via the activation of TRAF6-dependent signaling pathways. These results suggest that CD166 directs MM cell homing to the BM and promotes MM disease progression and osteolytic disease. CD166 may serve as a therapeutic target in the treatment of MM.Item Chloroquine increases osteoclast activity in vitro but does not improve the osteopetrotic bone phenotype of ADO2 mice(Elsevier, 2021) Alam, Imranul; Gerard-O’Riley, Rita L.; Acton, Dena; Hardman, Sara L.; Hong, Jung Min; Bruzzaniti, Angela; Econs, Michael J.; Medicine, School of MedicineAutosomal Dominant Osteopetrosis type II (ADO2) is a bone disease of impaired osteoclastic bone resorption that usually results from heterozygous missense mutations in the chloride channel 7 (CLCN7) gene. We created mouse models of ADO2 by introducing a knock-in (p.G213R) mutation in the Clcn7 gene, which is analogous to one of the common mutations (G215R) found in humans. The mutation leads to severe osteopetrosis and lethality in homozygous mice but produces substantial phenotypic variability in heterozygous mice on different genetic backgrounds that phenocopy the human disease of ADO2. ADO2 is an osteoclast-intrinsic disease, and lysosomal enzymes and proteins are critical for osteoclast activity. Chloroquine (CQ) is known to affect lysosomal trafficking, intracellular signaling and the lysosomal and vesicular pH, suggesting it might improve ADO2 osteoclast function. We tested this hypothesis in cell culture studies using osteoclasts derived from wild-type (WT or ADO2+/+) and ADO2 heterozygous (ADO2+/−) mice and found that CQ and its metabolite desethylchloroquine (DCQ), significantly increased ADO2+/− osteoclasts bone resorption activity in vitro, whereas bone resorption of ADO2+/+ osteoclasts was increased only by DCQ. In addition, we exploited our unique animal model of ADO2 on 129 background to identify the effect of CQ for the treatment of ADO2. Female ADO2 mice at 8 weeks of age were treated with 5 doses of CQ (1, 2.5, 5, 7.5 and 10 mg/kg BW/day) via drinking water for 6 months. Bone mineral density and bone micro-architecture were analyzed by longitudinal in-vivo DXA and micro-CT at baseline, 3 and 6 months. Serum bone biomarkers (CTX, TRAP and P1NP) were also analyzed at these time points. CQ treatment at the doses tested failed to produce any significant changes of aBMD, BMC (whole body, femur and spine) and trabecular BV/TV (distal femur) in ADO2 mice compared to the control group (water only). Further, levels of bone biomarkers were not significantly changed due to CQ treatment in these mice. Our findings indicate that while CQ increased osteoclast activity in vitro, it did not improve the osteopetrotic bone phenotypes in ADO2 heterozygous mice.Item CNC-bZIP protein NFE2L1 regulates osteoclast differentiation in antioxidant-dependent and independent manners(Elsevier, 2021-11-06) Liu, Zhiyuan; Wang, Huihui; Hou, Yongyong; Yang, Yang; Jia, Jingkun; Wu, Jinzhi; Zuo, Zhuo; Gao, Tianchang; Ren, Suping; Bian, Yiying; Liu, Shengnan; Fu, Jingqi; Sun, Yongxin; Li, Jiliang; Yamamoto, Masayuki; Zhang, Qiang; Xu, Yuanyuan; Pi, Jingbo; Biology, School of ScienceFine-tuning of osteoclast differentiation (OD) and bone remodeling is crucial for bone homeostasis. Dissecting the mechanisms regulating osteoclastogenesis is fundamental to understanding the pathogenesis of various bone disorders including osteoporosis and arthritis. Nuclear factor erythroid 2-related factor 1 (NFE2L1, also known as NRF1), which belongs to the CNC-bZIP family of transcription factors, orchestrates a variety of physiological processes and stress responses. While Nfe2l1 gene may be transcribed into multiple alternatively spliced isoforms, the biological function of the different isoforms of NFE2L1 in bone metabolism, osteoclastogenesis in particular, has not been reported. Here we demonstrate that knockout of all isoforms of Nfe2l1 transcripts specifically in the myeloid lineage in mice [Nfe2l1(M)-KO] results in increased activity of osteoclasts, decreased bone mass and worsening of osteoporosis induced by ovariectomy and aging. In comparison, LysM-Cre-mediated Nfe2l1 deletion has no significant effect on the osteoblast and osteocytes. Mechanistic investigations using bone marrow cells and RAW 264.7 cells revealed that deficiency of Nfe2l1 leads to accelerated and elevated OD, which is attributed, at least in part, to enhanced accumulation of ROS in the early stage of OD and expression of nuclear factor of activated T cells, cytoplasmic, calcineurin dependent 1α (Nfatc1/α). In addition, NFE2L1 regulates the transcription of multiple antioxidant genes and Nfatc1/α and OD in an isoform-specific manner. While long isoforms of NFE2L1 function as accelerators of induction of Nfatc1/α and antioxidant genes and OD, the short isoform NFE2L1-453 serves as a brake that keeps the long isoforms’ accelerator effects in check. These findings provide a novel insight into the regulatory roles of NFE2L1 in osteoclastogenesis and highlight that NFE2L1 is essential in regulating bone remodeling and thus may be a valuable therapeutic target for bone disorders.Item Crosstalk between dihydroceramides produced by Porphyromonas gingivalis and host lysosomal cathepsin B in the promotion of osteoclastogenesis(Wiley, 2022) Duarte, Carolina; Yamada, Chiaki; Garcia, Christopher; Akkaoui, Juliet; Ho, Anny; Nichols, Frank; Movila, Alexandru; Biomedical Sciences and Comprehensive Care, School of DentistryEmerging studies indicate that intracellular eukaryotic ceramide species directly activate cathepsin B (CatB), a lysosomal-cysteine-protease, in the cytoplasm of osteoclast precursors (OCPs) leading to elevated RANKL-mediated osteoclastogenesis and inflammatory osteolysis. However, the possible impact of CatB on osteoclastogenesis elevated by non-eukaryotic ceramides is largely unknown. It was reported that a novel class of phosphoglycerol dihydroceramide (PGDHC), produced by the key periodontal pathogen Porphyromonas gingivalis upregulated RANKL-mediated osteoclastogenesis in vitro and in vivo. Therefore, the aim of this study was to evaluate a crosstalk between host CatB and non-eukaryotic PGDHC on the promotion of osteoclastogenesis. According to a pulldown assay, high affinity between PGDHC and CatB was observed in RANKL-stimulated RAW264.7 cells in vitro. It was also demonstrated that PGDHC promotes enzymatic activity of recombinant CatB protein ex vivo and in RANKL-stimulated osteoclast precursors in vitro. Furthermore, no or little effect of PGDHC on the RANKL-primed osteoclastogenesis was observed in male and female CatB-knock out mice compared with their wild type counterparts. Altogether, these findings demonstrate that bacterial dihydroceramides produced by P. gingivalis elevate RANKL-primed osteoclastogenesis via direct activation of intracellular CatB in OCPs.Item Editorial: Bone inside-out and outside-in signals: Control of body homeostasis(Frontiers Media, 2023-01-06) Sankar, Uma; Brun, Lucas R.; Plotkin, Lilian I.; Anatomy, Cell Biology and Physiology, School of MedicineItem Editorial: Genetic and molecular determinants in bone health and diseases(Frontiers Media, 2024-01-17) Rossi, Michela; Lowery, Jonathan W.; Del Fattore, Andrea; Orthopaedic Surgery, School of MedicineItem Generation of the first Autosomal Dominant Osteopetrosis Type II (ADO2) disease models(Elsevier B.V., 2014-02) Alam, Imranul; Gray, Amie K.; Chu, Kang; Ichikawa, Shoji; Mohammad, Khalid S.; Capannolo, Marta; Capulli, Mattia; Maurizi, Antonio; Muraca, Maurizio; Teti, Anna; Econs, Michael J.; Del Fattore, Andrea; Department of Orthopaedic Surgery, IU School of MedicineAutosomal Dominant Osteopetrosis Type II (ADO2) is a heritable osteosclerotic disorder dependent on osteoclast impairment. In most patients it results from heterozygous missense mutations in the chloride channel 7 (CLCN7) gene, encoding for a 2Cl−/1H+ antiporter. By a knock-in strategy inserting a missense mutation in the Clcn7 gene, our two research groups independently generated mouse models of ADO2 on different genetic backgrounds carrying the homolog of the most frequent heterozygous mutation (p.G213R) in the Clcn7 gene found in humans. Our results demonstrate that the heterozygous model holds true presenting with higher bone mass, increased numbers of poorly resorbing osteoclasts and a lethal phenotype in the homozygous state. Considerable variability is observed in the heterozygous mice according with the mouse background, suggesting that modifier genes could influence the penetrance of the disease gene.Item Glucocorticoid Excess in Bone and Muscle(Springer, 2018-03) Sato, Amy Y.; Peacock, Munro; Bellido, Teresita; Anatomy and Cell Biology, School of MedicineGlucocorticoids (GC), produced and released by the adrenal glands, regulate numerous physiological processes in a wide range of tissues. Because of their profound immunosuppressive and anti-inflammatory actions, GC are extensively used for the treatment of immune and inflammatory conditions, the management of organ transplantation, and as a component of chemotherapy regimens for cancers. However, both pathologic endogenous elevation and long-term use of exogenous GC are associated with severe adverse effects. In particular, excess GC has devastating effects on the musculoskeletal system. GC increase bone resorption and decrease formation leading to bone loss, microarchitectural deterioration and fracture. GC also induce loss of muscle mass and strength leading to an increased incidence of falls. The combined effects on bone and muscle account for the increased fracture risk with GC. This review summarizes the advance in knowledge in the last two decades about the mechanisms of action of GC in bone and muscle and the attempts to interfere with the damaging actions of GC in these tissues with the goal of developing more effective therapeutic strategies.