scholarly journals A non-immunological role for γ-interferon–inducible lysosomal thiol reductase (GILT) in osteoclastic bone resorption

2021 ◽  
Vol 7 (17) ◽  
pp. eabd3684
Author(s):  
Benjamin W. Ewanchuk ◽  
Corey R. Arnold ◽  
Dale R. Balce ◽  
Priyatha Premnath ◽  
Tanis L. Orsetti ◽  
...  
Keyword(s):  
Bone Resorption ◽  
Cysteine Protease ◽  
Antigen Processing ◽  
Matrix Protein ◽  
Direct Reduction ◽  
Bone Matrix ◽  
Cathepsin K ◽  
Dependent Manner ◽  
Thiol Reductase ◽  
Γ Interferon

The extracellular bone resorbing lacuna of the osteoclast shares many characteristics with the degradative lysosome of antigen-presenting cells. γ-Interferon–inducible lysosomal thiol reductase (GILT) enhances antigen processing within lysosomes through direct reduction of antigen disulfides and maintenance of cysteine protease activity. In this study, we found the osteoclastogenic cytokine RANKL drove expression of GILT in osteoclast precursors in a STAT1-dependent manner, resulting in high levels of GILT in mature osteoclasts, which could be further augmented by γ-interferon. GILT colocalized with the collagen-degrading cysteine protease, cathepsin K, suggesting a role for GILT inside the osteoclastic resorption lacuna. GILT-deficient osteoclasts had reduced bone-resorbing capacity, resulting in impaired bone turnover and an osteopetrotic phenotype in GILT-deficient mice. We demonstrated that GILT could directly reduce the noncollagenous bone matrix protein SPARC, and additionally, enhance collagen degradation by cathepsin K. Together, this work describes a previously unidentified, non-immunological role for GILT in osteoclast-mediated bone resorption.

scholarly journals Inhibitory Effects of 2N1HIA (2-(3-(2-Fluoro-4-Methoxyphenyl)-6-Oxo-1(6H)-Pyridazinyl)-N-1H-Indol-5-Ylacetamide) on Osteoclast Differentiation via Suppressing Cathepsin K Expression

Molecules ◽  
2018 ◽  
Vol 23 (12) ◽  
pp. 3139 ◽  
Author(s):  
Sun-Hee Ahn ◽  
Zhihao Chen ◽  
Jinkyung Lee ◽  
Seok-Woo Lee ◽  
Sang Min ◽  
...  
Keyword(s):  
Bone Resorption ◽  
Osteoclast Differentiation ◽  
Cell Formation ◽  
Cathepsin K ◽  
Bone Diseases ◽  
Specific Gene ◽  
Tartrate Resistant Acid Phosphatase ◽  
Potential Candidate ◽  
Dependent Manner ◽  
Gene Markers

Osteoclasts are large multinucleated cells which are induced by the regulation of the receptor activator of nuclear factor kappa-Β ligand (RANKL), which is important in bone resorption. Excessive osteoclast differentiation can cause pathologic bone loss and destruction. Numerous studies have targeted molecules inhibiting RANKL signaling or bone resorption activity. In this study, 11 compounds from commercial libraries were examined for their effect on RANKL-induced osteoclast differentiation. Of these compounds, only 2-(3-(2-fluoro-4-methoxyphenyl)-6-oxo-1(6H)-pyridazinyl)-N-1H-indol-5-ylacetamide (2N1HIA) caused a significant decrease in multinucleated tartrate-resistant acid phosphatase (TRAP)-positive cell formation in a dose-dependent manner, without inducing cytotoxicity. The 2N1HIA compound neither affected the expression of osteoclast-specific gene markers such as TRAF6, NFATc1, RANK, OC-STAMP, and DC-STAMP, nor the RANKL signaling pathways, including p38, ERK, JNK, and NF-κB. However, 2N1HIA exhibited a significant impact on the expression levels of CD47 and cathepsin K, the early fusion marker and critical protease for bone resorption, respectively. The activity of matrix metalloprotease-9 (MMP-9) decreased due to 2N1HIA treatment. Accordingly, bone resorption activity and actin ring formation decreased in the presence of 2N1HIA. Taken together, 2N1HIA acts as an inhibitor of osteoclast differentiation by attenuating bone resorption activity and may serve as a potential candidate in preventing and/or treating osteoporosis, or other bone diseases associated with excessive bone resorption.

scholarly journals Suppressive effect of melatonin on osteoclast function via osteocyte calcitonin

2019 ◽  
Vol 242 (2) ◽  
pp. 13-23 ◽  
Author(s):  
Masaki Nakano ◽  
Mika Ikegame ◽  
Junko Igarashi-Migitaka ◽  
Yusuke Maruyama ◽  
Nobuo Suzuki ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Bone Matrix ◽  
Cathepsin K ◽  
Suppressive Effect ◽  
Melatonin Receptors ◽  
Dependent Manner ◽  
In Ovo ◽  
Expression Levels ◽  
Underlying Mechanisms ◽  
Mrna Expression Levels

Many studies have investigated the actions of melatonin on osteoblasts and osteoclasts. However, the underlying mechanisms, especially regarding osteocyte function, remain largely unknown. Therefore, this study aimed to clarify the underlying mechanisms of melatonin action on bone tissue via osteocyte function. Chick calvariae were employed as a model. In ovo injection of melatonin (5, 50 and 500 µg) dose-dependently decreased the mRNA expression levels of cathepsin K and matrix metalloproteinase 9 (MMP9) in chick calvariae without affecting the expression levels of receptor activator of NF-κB ligand or osteoprotegerin. Surprisingly enough, the expression of calcitonin mRNA in chick calvariae was significantly raised. After 3 days of in vitro treatment of melatonin (10−7 and 10−5 M) on newly hatched chick calvariae, both calcitonin mRNA expression in calvariae and the concentration of calcitonin in cultured medium were augmented in a dose-dependent manner, coincident with the decreased mRNA expression levels of cathepsin K and MMP9. Immunohistochemical analyses revealed expression of melatonin receptors and calcitonin by osteocytes buried in bone matrix. Moreover, the mRNA expression levels of melatonin receptors, calcitonin and sclerostin (a marker of osteocyte), were strongly and positively correlated. In conclusion, we demonstrated the expression of melatonin receptors and calcitonin expression in osteocytes for the first time and suggest a new mechanism underlying the suppressive effect of melatonin on osteoclasts via upregulation of calcitonin secretion by osteocytes.

Alpinetin Inhibits RANKL-Induced Osteoclastogenesis and Ovariectomy-Induced Bone Loss by Modulating NFATc1 Transcription and Lysosomal Function

2020 ◽  
Author(s):  
Rongxin He ◽  
Jinwei Lu ◽  
Yazhou Chen ◽  
Yong Li ◽  
Chenyi Ye ◽  
...  
Keyword(s):  
Bone Resorption ◽  
Bone Loss ◽  
Bone Matrix ◽  
Osteoclast Differentiation ◽  
Estrogen Deficiency ◽  
Bone Diseases ◽  
Dependent Manner ◽  
Metabolic Bone Diseases ◽  
Lysosomal Function ◽  
Metabolic Bone

Abstract BackgroundPostmenopausal osteoporosis is a chronic metabolic bone disease caused by excessive osteoclast activation, and osteoclasts are considered to be the sole participants in the degeneration and resorption of bone matrix for controlling bone integrity and continuity. The biological functions of osteoclasts depend critically on the number and activity of fused polykaryon. Hence, targeting osteoclast differentiation and activity can modulate bone resorption and alleviate osteoporosis. Alpinetin is widely used for excellent anti-inflammatory activities and little side-effect, but its role in osteoporosis remains unknown.ResultsIn this study, we investigated for the first time the ability of alpinetin to inhibit estrogen deficiency-induced bone loss. Alpinetin significantly reduced the expression levels of NFATc1 and its downstream genes, thereby inhibiting osteoclast differentiation in a concentration- and time-dependent manner. Additionally, alpinetin inhibited F-actin ring formation and bone resorption, as well as reduced the activation levels of NF-κB, ERK, and AKT signaling cascades. In mature osteoclasts, alpinetin remarkably inhibited integrin-mediated migration and lysosomal biogenesis and trafficking by modulating the PKCβ/TFEB and ATG5/LC3 axes. Importantly, alpinetin treatment in mice alleviated ovariectomy-induced bone volume loss. ConclusionOur findings strongly suggest that alpinetin plays a significant role in the regulation of NFATc1 production for the differentiation of osteoclasts and inhibits integrin-mediated cell migration and lysosomal function in mature osteoclasts, thus weaken the increased osteolytic ability due to estrogen deficiency. Alpinetin may represent a promising agent for the treatment of osteoporosis and other metabolic bone diseases.

scholarly journals N-[2-(4-Acetyl-1-Piperazinyl)Phenyl]-2-(3-Methylphenoxy)Acetamide (NAPMA) Inhibits Osteoclast Differentiation and Protects against Ovariectomy-Induced Osteoporosis

Molecules ◽  
2020 ◽  
Vol 25 (20) ◽  
pp. 4855
Author(s):  
Jinkyung Lee ◽  
Sun-Hee Ahn ◽  
Zhihao Chen ◽  
Sohi Kang ◽  
Dong Kyu Choi ◽  
...  
Keyword(s):  
Bone Resorption ◽  
Bone Loss ◽  
Osteoclast Differentiation ◽  
Cathepsin K ◽  
Bone Diseases ◽  
Drug Candidate ◽  
Tartrate Resistant Acid Phosphatase ◽  
Dependent Manner ◽  
Osteoclast Activity ◽  
Protein Levels

Osteoclasts are large, multinucleated cells responsible for bone resorption and are induced in response to the regulatory activity of receptor activator of nuclear factor-kappa B ligand (RANKL). Excessive osteoclast activity causes pathological bone loss and destruction. Many studies have investigated molecules that specifically inhibit osteoclast activity by blocking RANKL signaling or bone resorption. In recent years, we screened compounds from commercial libraries to identify molecules capable of inhibiting RANKL-induced osteoclast differentiation. Consequently, we reported some compounds that are effective at attenuating osteoclast activity. In this study, we found that N-[2-(4-acetyl-1-piperazinyl)phenyl]-2-(3-methylphenoxy)acetamide (NAPMA) significantly inhibited the formation of multinucleated tartrate-resistant acid phosphatase (TRAP)-positive cells from bone marrow-derived macrophages in a dose-dependent manner, without cytotoxic effects. NAPMA downregulated the expression of osteoclast-specific markers, such as c-Fos, NFATc1, DC-STAMP, cathepsin K, and MMP-9, at the transcript and protein levels. Accordingly, bone resorption and actin ring formation were decreased in response to NAPMA treatment. Furthermore, we demonstrated the protective effect of NAPMA against ovariectomy-induced bone loss using micro-CT and histological analysis. Collectively, the results showed that NAPMA inhibited osteoclast differentiation and attenuated bone resorption. It is thus a potential drug candidate for the treatment of osteoporosis and other bone diseases associated with excessive bone resorption.

scholarly journals Decreased C-Src Expression Enhances Osteoblast Differentiation and Bone Formation

2000 ◽  
Vol 151 (2) ◽  
pp. 311-320 ◽  
Author(s):  
Marilena Marzia ◽  
Natalie A. Sims ◽  
Susanne Voit ◽  
Silvia Migliaccio ◽  
Anna Taranta ◽  
...  
Keyword(s):  
Bone Resorption ◽  
Bone Formation ◽  
Osteoblast Differentiation ◽  
Matrix Protein ◽  
Bone Matrix ◽  
Osteogenic Cells ◽  
Alp Activity ◽  
Pthrp Receptor ◽  
Bone Matrix Protein

c-src deletion in mice leads to osteopetrosis as a result of reduced bone resorption due to an alteration of the osteoclast. We report that deletion/reduction of Src expression enhances osteoblast differentiation and bone formation, contributing to the increase in bone mass. Bone histomorphometry showed that bone formation was increased in Src null compared with wild-type mice. In vitro, alkaline phosphatase (ALP) activity and nodule mineralization were increased in primary calvarial cells and in SV40-immortalized osteoblasts from Src−/− relative to Src+/+ mice. Src-antisense oligodeoxynucleotides (AS-src) reduced Src levels by ∼60% and caused a similar increase in ALP activity and nodule mineralization in primary osteoblasts in vitro. Reduction in cell proliferation was observed in primary and immortalized Src−/− osteoblasts and in normal osteoblasts incubated with the AS-src. Semiquantitative reverse transcriptase-PCR revealed upregulation of ALP, Osf2/Cbfa1 transcription factor, PTH/PTHrP receptor, osteocalcin, and pro-alpha 2(I) collagen in Src-deficient osteoblasts. The expression of the bone matrix protein osteopontin remained unchanged. Based on these results, we conclude that the reduction of Src expression not only inhibits bone resorption, but also stimulates osteoblast differentiation and bone formation, suggesting that the osteogenic cells may contribute to the development of the osteopetrotic phenotype in Src-deficient mice.

scholarly journals Absence of γ-Interferon–inducible Lysosomal Thiol Reductase in Melanomas Disrupts T Cell Recognition of Select Immunodominant Epitopes

2002 ◽  
Vol 195 (10) ◽  
pp. 1267-1277 ◽  
Author(s):  
M. Azizul Haque ◽  
Ping Li ◽  
Sheila K. Jackson ◽  
Hassane M. Zarour ◽  
John W. Hawes ◽  
...  
Keyword(s):  
T Cell ◽  
Tumor Cell ◽  
T Cell Activation ◽  
Antigen Processing ◽  
Class Ii ◽  
Cell Recognition ◽  
T Cell Recognition ◽  
Thiol Reductase ◽  
Γ Interferon ◽  
Antigenic Epitopes

Long-lasting tumor immunity requires functional mobilization of CD8+ and CD4+ T lymphocytes. CD4+ T cell activation is enhanced by presentation of shed tumor antigens by professional antigen-presenting cells (APCs), coupled with display of similar antigenic epitopes by major histocompatibility complex class II on malignant cells. APCs readily processed and presented several self-antigens, yet T cell responses to these proteins were absent or reduced in the context of class II+ melanomas. T cell recognition of select exogenous and endogenous epitopes was dependent on tumor cell expression of γ-interferon–inducible lysosomal thiol reductase (GILT). The absence of GILT in melanomas altered antigen processing and the hierarchy of immunodominant epitope presentation. Mass spectral analysis also revealed GILT's ability to reduce cysteinylated epitopes. Such disparities in the profile of antigenic epitopes displayed by tumors and bystander APCs may contribute to tumor cell survival in the face of immunological defenses.

scholarly journals DIHYDROTESTOSTERONE DOWNREGULATES BONE RESORPTION ACTIVITY OF OSTEOCLASTS IN DOSE DEPENDENT MANNER: AN IN VITRO MODEL USING RAW 264.7 CELLS

2017 ◽  
Vol 9 (10) ◽  
pp. 86
Author(s):  
Hnin Ei Thu ◽  
Zahid Hussain ◽  
Isa Naina Mohamed ◽  
Ahmad Nazrun Shuid
Keyword(s):  
Bone Resorption ◽  
Cathepsin K ◽  
Raw 264.7 Cells ◽  
Tartrate Resistant Acid Phosphatase ◽  
Raw 264.7 ◽  
Dependent Manner ◽  
Sod Activity ◽  
Osteoclastic Differentiation ◽  
Dose Dependent

Objective: Numerous studies have evidenced the bone regulatory potential of dihydrotestosterone in androgen-deficient osteoporosis. The present study was thus aimed to explore the translational mechanism of dihydrotestosterone to down-regulate the bone resorption activity of osteoclasts using RAW 264.7 cells as in vitro model.Methods: Prior to analyze the efficacy of dihydrotestosterone (5α-DHT) to alleviate osteoclastic differentiation, their cell viability and cell proliferative ability was assessed using lactate dehydrogenase (LDH) and MTS assays. The osteoclastic differentiation capacity of dihydrotestosterone was evaluated by measuring TRAP activity and the expression of bone resorption-related proteins such as matrix metallopeptidase-9 (MMP-9), cathepsin-K, tartrate-resistant acid phosphatase (TRAP) and NFATc1. Moreover, the effects of dihydrotestosterone were also evaluated on superoxide (free radicals) generation and superoxide dismutase (SOD) activity in RANKL-induced osteoclasts.Results: Dihydrotestosterone showed no toxicity towards RAW 264.7 cells and significantly enhanced their proliferation and growth rates in a dose-dependent fashion. It was also observed that dihydrotestosterone exhibits a remarkable inhibitory effect on differentiation, maturation and activation of osteoclasts. The marked inhibition of differentiation and activation of osteoclasts caused by 5α-DHT was due to down-regulation of the expression of MMP-9, cathepsin-K, TRAP, NFATc1, generation of superoxide and up-regulation of SOD activity in the RAW 264.7 cells.Conclusion: Resulting data provided substantially in vitroevidence for the pronounced anti-osteoclastogenetic activity of dihydrotestosterone and its therapeutic value in treating osteoporosis and other bone-erosive disorders. 

Localization of Rat Cathepsin K in Osteoclasts and Resorption Pits: Inhibition of Bone Resorption and Cathepsin K-Activity by Peptidyl Vinyl Sulfones

1999 ◽  
Vol 380 (6) ◽  
Author(s):  
L. Xia ◽  
J. Kilb ◽  
H. Wex ◽  
Z. Li ◽  
A. Lipyansky ◽  
...  
Keyword(s):  
Bone Resorption ◽  
Cathepsin L ◽  
Critical Role ◽  
Specific Inhibitor ◽  
Cathepsin K ◽  
Selective Inhibition ◽  
Specific Substrate ◽  
Dependent Manner ◽  
Protease Inhibition ◽  
K Activity

AbstractWe have localized cathepsin K in rat osteoclasts and within exposed resorption pits by immuno-fluorescence microscopy. Intracellular staining using an antibody raised against recombinant mouse cathepsin K was vesicular and uniformly distributed throughout the cell. Confocal microscopy analysis did not reveal an accumulation of cathepsin K containing vesicles opposing the ruffled border and the resorption lacuna. Exposed resorption pits exhibited a uniform distribution of cathepsin K, and no differences were observed between the edges and the centers of the pits. The immunostaining of resorption pits with anti-cathepsin K antibodies demonstrates that the protease is secreted into the sub-osteoclastic compartment.Cathepsin K-specific inhibition using peptidyl vinyl sulfones as selective cysteine protease inactivators reduced bone resorption by 80% in a dose-dependent manner at sub-micromolar concentrations. No reduction of bone resorption was observed at those low concentrations using a potent cathepsin L, S, B-specific inhibitor. That the inhibition of bone resorption can be attributed to cathepsin K-like protease inhibition was corroborated by the selective inhibition of the osteoclastic Z-Gly-Pro-Arg-MβNA hydrolyzing activity by the cathepsin K, L, S, B-inhibitor, but not by the cathepsin L, B, and S inhibitor. Z-Gly-Pro-Arg-MβNA is efficiently hydrolyzed by cathepsin K but only poorly by cathepsins L, S, and B. On the contrary, the intracellular hydrolysis of the cathepsin B-specific substrate, Z-Arg-Arg-MβNA, was prevented by both types of inhibitors.The identification of cathepsin K in resorption pits and the inhibition of bone resorption and intracellular cathepsin K activity by selective vinyl sulfone inhibitors indicate the critical role of the protease in osteoclastic bone resorption.

Endocrine function of osteocytes

2016 ◽  
Vol 25 (02) ◽  
pp. 62-68
Author(s):  
T. Yorgan ◽  
T. Schinke
Keyword(s):  
Bone Formation ◽  
Matrix Protein ◽  
Vascular System ◽  
Bone Matrix ◽  
Endocrine Function ◽  
Whole Body ◽  
Dependent Manner ◽  
Phosphate Homeostasis ◽  
Target Organs ◽  
Additional Influence

SummaryOsteocytes represent the most abundant cell type of the skeletal system. They have access to a large cellular surface area within the lacuno-canalicular network. This network additionally provides connection to the vascular system, a prerequisite for secretion of endocrine regulators into the circulation. The best established endocrine function of the osteocyte network is the regulation of phosphate homeostasis by secretion of Fgf23, a hormone inhibiting renal phosphate reabsorption. Recently, several additional osteocyte-derived factors have been suggested to influence phosphate homeostasis, either directly or in an Fgf23-dependent manner. Moreover, osteocytes are also the major producers of Wnt signaling modulators, such as Sclerostin or Dkk1. Since these molecules primarily act as inhibitors of bone formation, there might be an additional influence of osteocyte-derived molecules on glucose handling and energy metabolism. In fact, osteocalcin, a long-known bone matrix protein and biomarker of bone formation, is now considered to act as a hormone controlling insulin production by pancreatic β-cells and insulin sensitivity of target organs. Since the endocrine functions of osteocytes are only beginning to be uncovered, it appears likely that additional osteocyte-derived molecules with systemic influences on whole body homeostasis might be identified in the future.

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