scholarly journals Rebamipide, an Amino Acid Analog of 2(1H)-Quinolinone, Inhibits the Formation of Human Osteoclasts

2016 ◽  
Vol 2016 ◽  
pp. 1-7 ◽  
Author(s):  
Yuki Nanke ◽  
Tsuyoshi Kobashigawa ◽  
Toru Yago ◽  
Manabu Kawamoto ◽  
Hisashi Yamanaka ◽  
...  
Keyword(s):  
Mononuclear Cells ◽  
Drug Repositioning ◽  
Transmembrane Protein ◽  
Time Lapse ◽  
Osteoclast Formation ◽  
Toxicity Tests ◽  
Peripheral Blood Mononuclear ◽  
Traditional Drug ◽  
Amino Acid Analog ◽  
Immunohistological Staining

Objectives. Drug repositioning or drug reprofiling (DR) has recently been growing in importance. DR has a significant advantage over traditional drug development because the repositioned drug has already passed toxicity tests; its safety is known, and the risk of adverse toxicology is reduced. In the current study, we investigated the role of rebamipide, a mucosa-protecting agent, with recently reported anti-inflammatory function, in human osteoclastogenesis. Methods. Peripheral blood mononuclear cells (PBMCs) were cultured in the presence of M-CSF and sRANKL. Osteoclast formation was evaluated by immunohistological staining for CD51/61 (vitronectin receptors). Osteoclast formation, in the presence or absence of rebamipide (0, 1, and 3 mM), was observed by time-lapse photography and actin ring formation. The number of absorption sites and area of absorption were calculated using Osteologic™ plates. Pit formation was studied by 3D-SEM. Results. Rebamipide inhibited human osteoclast formation at 3 mM, a pharmacological concentration, and inhibited resorbing activity dose-dependently. Rebamipide induced the degradation of actin rings in mature osteoclasts. This mechanism may involve inhibiting the osteoclast fusion pathway through reducing the expression of DC-specific transmembrane protein (DC-STAMP). Conclusions. The present study suggests that rebamipide would be useful as a novel agent for osteoporosis and rheumatoid arthritis.

Cell-cell fusion: human multinucleated osteoclasts

2009 ◽  
Vol 4 (4) ◽  
pp. 543-548 ◽  
Author(s):  
Zhi-Yong Zeng ◽  
Jun-Min Chen
Keyword(s):  
Cell Fusion ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Transmembrane Protein ◽  
Osteoclast Differentiation ◽  
Osteoclast Formation ◽  
Tartrate Resistant Acid Phosphatase ◽  
Peripheral Blood Mononuclear ◽  
Healthy Donors ◽  
Cell Cell

AbstractOsteoclasts are known to be formed by fusion of circulating mononuclear precursor cells which originate from haematopoietic stem cells. The precise mechanisms regulating the cell-cell fusion of these circulating cells to multinucleated osteoclasts remain unclear. In the present study, human peripheral blood mononuclear cells (PBMNCs) from healthy donors were treated with the macrophagecolony stimulating factor (M-CSF) and receptor activator of nuclear factor (NF)-κB ligand (RANKL) to induce osteoclast differentiation. Osteoclast formation and resorption activity were investigated through the use of tartrate-resistant acid phosphatase (TRAP) staining and lacunar resorption on dentine slices respectively. Real-time reverse-transcription polymerase chain reaction (PCR) was used to detect expression of dendritic cell-specific transmembrane protein (DC-STAMP) in these cells. The results showed that under the treatment of M-CSF and RANKL, PBMNCs differentiated into multinucleated osteoclasts through cell-cell fusion of mononucleated cells. These osteoclasts were TRAP positive and capable of resorbing the bone. Expression of DC-STAMP was much higher in the cells treated with both M-CSF and RANKL than those treated with M-CSF alone. We concluded that human PBMNCs might differentiate into active osteoclasts under certain conditions and the DC-STAMP, which is believed critical for osteoclast development, will be a possible therapeutic target for osteoclast related diseases in future.

scholarly journals Cells Derived from Human Long Bone Appear More Differentiated and More Actively Stimulate Osteoclastogenesis Compared to Alveolar Bone-Derived Cells

2020 ◽  
Vol 21 (14) ◽  
pp. 5072
Author(s):  
Cindy Kelder ◽  
Cornelis J. Kleverlaan ◽  
Marjolijn Gilijamse ◽  
Astrid D. Bakker ◽  
Teun J. de Vries
Keyword(s):  
Tissue Engineering ◽  
Mononuclear Cells ◽  
Alveolar Bone ◽  
Bone Cells ◽  
Transmembrane Protein ◽  
Cell Types ◽  
Long Bone ◽  
Long Bones ◽  
Peripheral Blood Mononuclear ◽  
Alveolar Bone Cells

Osteoblasts derived from mouse skulls have increased osteoclastogenic potential compared to long bone osteoblasts when stimulated with 1,25(OH)2 vitamin D3 (vitD3). This indicates that bone cells from specific sites can react differently to biochemical signals, e.g., during inflammation or as emitted by bioactive bone tissue-engineering constructs. Given the high turn-over of alveolar bone, we hypothesized that human alveolar bone-derived osteoblasts have an increased osteogenic and osteoclastogenic potential compared to the osteoblasts derived from long bone. The osteogenic and osteoclastogenic capacity of alveolar bone cells and long bone cells were assessed in the presence and absence of osteotropic agent vitD3. Both cell types were studied in osteogenesis experiments, using an osteogenic medium, and in osteoclastogenesis experiments by co-culturing osteoblasts with peripheral blood mononuclear cells (PBMCs). Both osteogenic and osteoclastic markers were measured. At day 0, long bones seem to have a more late-osteoblastic/preosteocyte-like phenotype compared to the alveolar bone cells as shown by slower proliferation, the higher expression of the matrix molecule Osteopontin (OPN) and the osteocyte-enriched cytoskeletal component Actin alpha 1 (ACTA1). This phenotype was maintained during the osteogenesis assays, where long bone-derived cells still expressed more OPN and ACTA1. Under co-culture conditions with PBMCs, long bone cells also had a higher Tumor necrose factor-alfa (TNF-α) expression and induced the formation of osteoclasts more than alveolar bone cells. Correspondingly, the expression of osteoclast genes dendritic cell specific transmembrane protein (DC-STAMP) and Receptor activator of nuclear factor kappa-Β ligand (RankL) was higher in long bone co-cultures. Together, our results indicate that long bone-derived osteoblasts are more active in bone-remodeling processes, especially in osteoclastogenesis, than alveolar bone-derived cells. This indicates that tissue-engineering solutions need to be specifically designed for the site of application, such as defects in long bones vs. the regeneration of alveolar bone after severe periodontitis.

Isopentenyl Pyrophosphate (IPP), a Metabolite produced in Myeloma Cells Induces the Chemotaxis of γδT Cells

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2766-2766
Author(s):  
Eishi Ashihara ◽  
Tatsuya Munaka ◽  
Shinya Kimura ◽  
Masaki Kanai ◽  
Hirohisa Abe ◽  
...  
Keyword(s):  
T Cells ◽  
Mononuclear Cells ◽  
Mevalonate Pathway ◽  
Ex Vivo ◽  
Time Lapse ◽  
Dependent Manner ◽  
Peripheral Blood Mononuclear ◽  
Isopentenyl Pyrophosphate ◽  
Γδt Cells ◽  
Cellular Analysis

Abstract Abstract 2766 γδT cells, which control the innate immune system, are classified into three subtypes on the basis of Vγ chain. Of these subtypes, Vγ2Vδ9 T cells display anti-tumor immunity. We have demonstrated that nitrogen-containing bisphosphonate (N-BP) treatment expands Vγ2Vδ9 T cells ex vivo and that these expanded cells can kill tumor cells in a major histocompatibility complex-unrestricted manner (Sato, Int J Cancer, 2005; Uchida, Biochem Biophys Res Commun, 2007; Sato, Cancer Immunol Immunother, 2008.). N-BP inhibits farnesyl pyrophosphate synthase in the mevalonate pathway, resulting in the accumulation of isopentenyl pyrophosphate (IPP), which is a stimulatory antigen for Vγ2Vδ9T cells. In the present study, we investigated the chemotactic factors for Vγ2Vδ9T cells by using a micro total analysis system-based microfluidic cellular analysis device (Kanai, Sens Actuators A, 2004; Munaka, Analyst, 2007.). This microchip possesses a minute-volume (240 nL) chamber integrated with a micro-sample injector that permits the injection of a small amount (several nL) of a solute (Figure 1). Because of the minute size of this chamber, a concentration gradient can be maintained free from the influence of fluid convection and stirring, and the solute can consequently spread in a diffusion-dependent manner. Therefore, administration of a humoral factor via the sample injector mimics its release from the cell surface. We first investigated whether the supernatant of RPMI8226 multiple myeloma (MM) cells treated with zoledronic acid (ZOL) induced chemotaxis of γδT cells. We treated RPMI8226 MM cells with ZOL (1 mM) overnight and collected the supernatant. Human γδT cells were obtained by the culture of peripheral blood mononuclear cells as previously reported (Uchida, Biochem Biophys Res Commun, 2007.), and these cells were cultured in the microchip. After the injection of supernatant, γδT cells migration was observed under a microscope and continuous time-lapse recording was performed for 30 min. γδT cells migrated toward the injector, indicating that the supernatant of ZOL-treated RPMI8226 cells includes a chemoattractant factor for γδT cells. We next applied soluble MICA (sMICA), sICAM-1, sVCAM-1, and IPP and examined the migration of γδT cells. Among them, sMICA and IPP were chemoattractive for γδT cells, and the velocity of γδT cell migration was increased by the injection of IPP compared to the solvent control (Figure 2). These observations indicate that IPP, a metabolite of the mevalonate pathway in MM cells, or sMICA is a chemotactic factor for γδT cells when the target MM ells are treated with ZOL. Disclosures: Munaka: Shimadzu Corporation: Employment. Kanai:Shimadzu Corporation: Employment. Abe:Shimadzu Corporation: Employment.

scholarly journals Evaluation of the Efficacy and Safety of a Clinical Grade Human Induced Pluripotent Stem Cell-Derived Cardiomyocyte Patch: A Pre-Clinical Study

2021 ◽  
Author(s):  
Shigeru Miyagawa ◽  
Takuji Kawamura ◽  
Emiko Ito ◽  
Maki Takeda ◽  
Hiroko Iseoka ◽  
...  
Keyword(s):  
Mononuclear Cells ◽  
Interstitial Fibrosis ◽  
Toxicity Tests ◽  
Sequencing Analysis ◽  
Clinical Grade ◽  
Efficacy And Safety ◽  
Peripheral Blood Mononuclear ◽  
Induced Pluripotent

Aims: Cardiomyocyte-derived induced pluripotent stem cells (iPSCs) may represent a promising therapeutic strategy for severely damaged myocardium. This study aimed to assess the efficacy and safety of clinical grade human iPSC-derived cardiomyocyte (hiPSC-CM) patches and conduct a pre-clinical proof-of-concept analysis. Methods and results: A clinical grade hiPSC line was established from peripheral blood mononuclear cells collected from a healthy volunteer homozygous for human leukocyte antigens and differentiated into cardiomyocytes using cytokines and chemical compounds. hiPSC-CMs were cultured on temperature-responsive culture dishes to fabricate the hiPSC-CM patch. The hiPSC-CMs expressed cardiomyocyte-specific genes and proteins while electrophysiological analyses revealed that hiPSC-CMs were similar to the human myocardium. In vitro safety studies using cell growth, soft agar colony formation, and undifferentiated cell assays indicated that tumourigenic cells were not present. Moreover, no genomic mutations were discovered using whole genome and exome sequencing analysis. Tumour formation was not detected in an in vivo tumourigenicity assay using NOG mice. General toxicity tests also showed no adverse events due to hiPSC-CM patch transplantation. An efficacy study using a porcine model of myocardial infarction demonstrated significantly improved cardiac function with angiogenesis and a reduction in interstitial fibrosis, which was enhanced by cytokine secretion from hiPSC-CM patches after transplantation. No lethal arrhythmias were observed. Conclusion: hiPSC-CM patches show promise for future translational research and clinical trials for ischaemic heart failure.

scholarly journals BCPA {N,N′-1,4-Butanediylbis[3-(2-chlorophenyl)acrylamide]} Inhibits Osteoclast Differentiation through Increased Retention of Peptidyl-Prolyl cis-trans Isomerase Never in Mitosis A-Interacting 1

2018 ◽  
Vol 19 (11) ◽  
pp. 3436 ◽  
Author(s):  
Eugene Cho ◽  
Jin-Kyung Lee ◽  
Jee-Young Lee ◽  
Zhihao Chen ◽  
Sun-Hee Ahn ◽  
...  
Keyword(s):  
Mononuclear Cells ◽  
Transmembrane Protein ◽  
Osteoclast Differentiation ◽  
Osteoclast Formation ◽  
Tartrate Resistant Acid Phosphatase ◽  
Mrna Levels ◽  
Dependent Manner ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
And Function

Osteoporosis is caused by an imbalance of osteoclast and osteoblast activities and it is characterized by enhanced osteoclast formation and function. Peptidyl-prolyl cis-trans isomerase never in mitosis A (NIMA)-interacting 1 (Pin1) is a key mediator of osteoclast cell-cell fusion via suppression of the dendritic cell-specific transmembrane protein (DC-STAMP). We found that N,N′-1,4-butanediylbis[3-(2-chlorophenyl)acrylamide] (BCPA) inhibited receptor activator of nuclear factor kappa-B ligand (RANKL)-induced osteoclastogenesis in a dose-dependent manner without cytotoxicity. In addition, BCPA attenuated the reduction of Pin1 protein during osteoclast differentiation without changing Pin1 mRNA levels. BCPA repressed the expression of osteoclast-related genes, such as DC-STAMP and osteoclast-associated receptor (OSCAR), without altering the mRNA expression of nuclear factor of activated T cells (NFATc1) and cellular oncogene fos (c-Fos). Furthermore, Tartrate-resistant acid phosphatase (TRAP)-positive mononuclear cells were significantly decreased by BCPA treatment compared to treatment with the Pin1 inhibitor juglone. These data suggest that BCPA can inhibit osteoclastogenesis by regulating the expression of the DC-STAMP osteoclast fusion protein by attenuating Pin1 reduction. Therefore, BCPA may be used to treat osteoporosis.

scholarly journals A Drug Repositioning Approach Identifies a Combination of Compounds as a Potential Regimen for Chronic Lymphocytic Leukemia Treatment

2021 ◽  
Vol 11 ◽  
Author(s):  
Atef Nehdi ◽  
Nosaibah Samman ◽  
Abdullah Mashhour ◽  
Alshaimaa Alhallaj ◽  
Thadeo Trivilegio ◽  
...  
Keyword(s):  
Mononuclear Cells ◽  
Kinase Inhibitor ◽  
Drug Repositioning ◽  
Human Peripheral Blood ◽  
Lymphocytic Leukemia ◽  
Atp Depletion ◽  
Leukemic Cells ◽  
Peripheral Blood Mononuclear ◽  
Intracellular Atp ◽  
Leukemia Treatment

Drug repositioning is a promising and powerful innovative strategy in the field of drug discovery. In this study, we screened a compound-library containing 800 Food and Drug Administration approved drugs for their anti-leukemic effect. All screening activities made use of human peripheral blood mononuclear cells (PBMCs), isolated from healthy or leukemic donors. Compounds with confirmed cytotoxicity were selected and classified in three groups: i) anti-neoplastic compounds which are drugs used in leukemia treatment, ii) compounds known to have an anti-cancer effect and iii) compounds demonstrating an anti-leukemic potential for the first time. The latter group was the most interesting from a drug repositioning perspective and yielded a single compound, namely Isoprenaline which is a non-selective β-adrenergic agonist. Analysis of the cytotoxic effect of this drug indicated that it induces sustainable intracellular ATP depletion leading, over time, to necrotic cell death. We exploited the Isoprenaline-induced intracellular ATP depletion to sensitize primary leukemic cells to fludarabine (purine analogue) and Ibrutinib (Bruton’s tyrosine kinase inhibitor) treatment. In-vitro treatment of primary leukemic cells with a combination of Isoprenaline/fludarabine or Isoprenaline/Ibrutinib showed a very high synergistic effect. These combinations could constitute a new efficient regimen for CLL treatment following successful evaluation in animal models and clinical trials.

scholarly journals Reduced miR-144-3p expression in serum and bone mediates osteoporosis pathogenesis by targeting RANK

2018 ◽  
Vol 96 (5) ◽  
pp. 627-635 ◽  
Author(s):  
Chunqing Wang ◽  
Hanliang He ◽  
Liang Wang ◽  
Yu Jiang ◽  
Youjia Xu
Keyword(s):  
Mononuclear Cells ◽  
Osteoclast Differentiation ◽  
Cathepsin K ◽  
Osteoclast Formation ◽  
Bone Homeostasis ◽  
Serum Samples ◽  
Peripheral Blood Mononuclear ◽  
Altered Expression ◽  
Rna Molecules ◽  
Non Coding Rna

Osteoblasts and osteoclasts are responsible for the formation and resorption of bone, respectively. An imbalance between these two processes results in a disease called osteoporosis, in which a decreased level of bone strength increases the risk of a bone fracture. MicroRNAs (miRNAs) are small non-coding RNA molecules of 18–25 nucleotides that have been previously shown to control bone metabolism by regulating osteoblast and osteoclast differentiation. In this study, we detected the expression pattern of 10 miRNAs in serum samples from patients with osteoporosis, and identified the altered expression of 6 miRNAs by comparison with patients without osteoporosis. We selected miR-144-3p for further investigation, and showed that it regulates osteoclastogenesis by targeting RANK, and that it is conserved amongst vertebrates. Disrupted expression of miR-144-3p in CD14+ peripheral blood mononuclear cells changed TRAP activity and the osteoclast-specific genes TRAP, cathepsin K (CTSK), and NFATC. TRAP staining, CCK-8, and flow cytometry analyses revealed that miR-144-3p also affects osteoclast formation, proliferation, and apoptosis. Together, these results indicate that miR-144-3p critically mediates bone homeostasis, and thus, represents a promising novel therapeutic candidate for the treatment of this disease.

scholarly journals I-BET151 suppresses osteoclast formation and inflammatory cytokines secretion by targetting BRD4 in multiple myeloma

2019 ◽  
Vol 39 (5) ◽  
Author(s):  
Ning-Hong Guo ◽  
Ji-Fu Zheng ◽  
Fu-Ming Zi ◽  
Jing Cheng
Keyword(s):  
Inflammatory Cytokines ◽  
Inflammatory Cytokine ◽  
Mononuclear Cells ◽  
Osteoclast Differentiation ◽  
Osteoclast Formation ◽  
Cytokine Secretion ◽  
Raw 264.7 ◽  
Peripheral Blood Mononuclear ◽  
Tnf Α ◽  
Brd4 Inhibition

Abstract Background: Multiple myeloma (MM) is an incurable hematologic cancer, accompanied by excessive osteoclast formation and inflammatory cytokine secretion. The mechanisms by which bromodomain and extra-terminal domain (BET) protein inhibitor I-BET151 regulates osteoclast differentiation and inflammatory cytokine secretion in MM are largely unknown. Methods: The isolated peripheral blood mononuclear cells from normal or patients with MM were treated with receptor activator of NF-κB ligand (RANKL) and M-CSF to induce osteoclast differentiation. RAW 264.7 cells were treated with RANKL. I-BET151 was applied to investigate the effects of BRD4 inhibition on osteoclast formation and inflammatory cytokine secretion. Osteoclast formation was determined by tartrate-resistant acid phosphatase (TRACP) staining. The expression of osteoclast-specific genes TRACP, matrix metalloproteinase-9 (MMP-9), cathepsin K (Ctsk), and c-Src was tested using quantitative real-time PCR. And the level of inflammatory cytokines TNF-α, IL-1β, and IL-6 was assessed by ELISA. Tumor necrosis factor receptor-associated factor 6 (TRAF6), BRD4, nuclear and cytoplasm p65, IκB-α, nuclear factor of activated T cells cytoplasmic (NFATc1), and osteoprotegerin (OPG) expression were measured by Western blotting. RNAi technology was applied to knock down BET family member BRD4. Results: I-BET151 dose-dependently suppressed osteoclast formation, inhibited the levels of osteoclast-specific genes TRACP, MMP-9, Ctsk, and c-Src and inflammatory cytokines TNF-α, IL-1β, and IL-6 secretion in peripheral blood mononuclear cells and RAW 264.7. I-BET151 inhibited the protein levels of BRD4 and NFATc1, increased OPG expression, and suppressed IκB-α degradation and p65 nuclear translocation. Further, the effects of I-BET151 on osteoclast formation, osteoclast-specific genes expression, inflammatory cytokine secretion, and NF-κB inhibition were promoted by BRD4 knockdown. Conclusion: I-BET151 inhibits osteoclast formation and inflammatory cytokine secretion by targetting BRD4-mediated RANKL-NF-κB signal pathway and BRD4 inhibition might be beneficial for MM treatment.

scholarly journals Prostaglandin E2 Strongly Inhibits Human Osteoclast Formation

Endocrinology ◽  
2005 ◽  
Vol 146 (12) ◽  
pp. 5204-5214 ◽  
Author(s):  
Ikuko Take ◽  
Yasuhiro Kobayashi ◽  
Yohei Yamamoto ◽  
Hideki Tsuboi ◽  
Takahiro Ochi ◽  
...  
Keyword(s):  
Cell Cultures ◽  
Peripheral Blood Mononuclear Cells ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Inhibitory Effect ◽  
Osteoclast Formation ◽  
Mouse Macrophage ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells ◽  
Human Osteoclast

Prostaglandin E2 (PGE2) enhances osteoclast formation in mouse macrophage cultures treated with receptor activator of nuclear factor-κB ligand (RANKL). The effects of PGE2 on human osteoclast formation were examined in cultures of CD14+ cells prepared from human peripheral blood mononuclear cells. CD14+ cells differentiated into osteoclasts in the presence of RANKL and macrophage colony-stimulating factor. CD14+ cells expressed EP2 and EP4, but not EP1 or EP3, whereas CD14+ cell-derived osteoclasts expressed none of the PGE2 receptors. PGE2 and PGE1 alcohol (an EP2/4 agonist) stimulated cAMP production in CD14+ cells. In contrast to mouse macrophage cultures, PGE2 and PGE1 alcohol inhibited RANKL-induced human osteoclast formation in CD14+ cell cultures. H-89 blocked the inhibitory effect of PGE2 on human osteoclast formation. These results suggest that the inhibitory effect of PGE2 on human osteoclast formation is mediated by EP2/EP4 signals. SaOS4/3 cells have been shown to support human osteoclast formation in cocultures with human peripheral blood mononuclear cells in response to PTH. PGE2 inhibited PTH-induced osteoclast formation in cocultures of SaOS4/3 cells and CD14+ cells. Conversely, NS398 (a cyclooxygenase 2 inhibitor) enhanced osteoclast formation induced by PTH in the cocultures. The conditioned medium of CD14+ cells pretreated with PGE2 inhibited RANKL-induced osteoclast formation not only in human CD14+ cell cultures, but also in mouse macrophage cultures. These results suggest that PGE2 inhibits human osteoclast formation through the production of an inhibitory factor(s) for osteoclastogenesis of osteoclast precursors.

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