Inhibition of lipopolysaccharide-induced osteoclast formation and bone resorption in vitro and in vivo by cysteine proteinase inhibitors

Although the anti-tumor and anti-infective properties of β-glucans have been well-discussed, their role in bone metabolism has not been reviewed so far. This review discusses the biological effects of β-glucans on bone metabolisms, especially on bone-resorbing osteoclasts, which are differentiated from hematopoietic precursors. Multiple immunoreceptors that can recognize β-glucans were reported to be expressed in osteoclast precursors. Coordinated co-stimulatory signals mediated by these immunoreceptors are important for the regulation of osteoclastogenesis and bone remodeling. Curdlan from the bacterium Alcaligenes faecalis negatively regulates osteoclast differentiation in vitro by affecting both the osteoclast precursors and osteoclast-supporting cells. We also showed that laminarin, lichenan, and glucan from baker’s yeast, as well as β-1,3-glucan from Euglema gracilisas, inhibit the osteoclast formation in bone marrow cells. Consistent with these findings, systemic and local administration of β-glucan derived from Aureobasidium pullulans and Saccharomyces cerevisiae suppressed bone resorption in vivo. However, zymosan derived from S. cerevisiae stimulated the bone resorption activity and is widely used to induce arthritis in animal models. Additional research concerning the relationship between the molecular structure of β-glucan and its effect on osteoclastic bone resorption will be beneficial for the development of novel treatment strategies for bone-related diseases.

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Nirogacestat suppresses RANKL-Induced osteoclast formation in vitro and attenuates LPS-Induced bone resorption in vivo

Experimental Cell Research ◽

10.1016/j.yexcr.2019.06.015 ◽

2019 ◽

Vol 382 (1) ◽

pp. 111470 ◽

Cited By ~ 4

Author(s):

Xuzhuo Chen ◽

Xinwei Chen ◽

Zhihang Zhou ◽

Yi Mao ◽

Yexin Wang ◽

...

Keyword(s):

Bone Resorption ◽

Osteoclast Formation

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Dihydroartemisinin prevents breast cancer-induced osteolysis via inhibiting both breast caner cells and osteoclasts

Scientific Reports ◽

10.1038/srep19074 ◽

2016 ◽

Vol 6 (1) ◽

Cited By ~ 20

Author(s):

Ming-Xuan Feng ◽

Jian-Xin Hong ◽

Qiang Wang ◽

Yong-Yong Fan ◽

Chi-Ting Yuan ◽

...

Keyword(s):

Breast Cancer ◽

Bone Resorption ◽

Molecular Mechanisms ◽

Osteoclast Differentiation ◽

Metastatic Breast ◽

Osteoclast Formation ◽

Migration And Invasion ◽

Titanium Particle

Abstract Bone is the most common site of distant relapse in breast cancer, leading to severe complications which dramatically affect the patients’ quality of life. It is believed that the crosstalk between metastatic breast cancer cells and osteoclasts is critical for breast cancer-induced osteolysis. In this study, the effects of dihydroartemisinin (DHA) on osteoclast formation, bone resorption, osteoblast differentiation and mineralization were initially assessed in vitro, followed by further investigation in a titanium-particle-induced osteolysis model in vivo. Based on the proved inhibitory effect of DHA on osteolysis, DHA was further applied to MDA-MB-231 breast cancer-induced mouse osteolysis model, with the underlying molecular mechanisms further investigated. Here, we verified for the first time that DHA suppressed osteoclast differentiation, F-actin ring formation and bone resorption through suppressing AKT/SRC pathways, leading to the preventive effect of DHA on titanium-particle-induced osteolysis without affecting osteoblast function. More importantly, we demonstrated that DHA inhibited breast tumor-induced osteolysis through inhibiting the proliferation, migration and invasion of MDA-MB-231 cells via modulating AKT signaling pathway. In conclusion, DHA effectively inhibited osteoclastogenesis and prevented breast cancer-induced osteolysis.

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Binding of FAD to 6-hydroxy-d-nicotine oxidase apoenzyme prevents degradation of the holoenzyme

Biochemical Journal ◽

10.1042/bj2580187 ◽

1989 ◽

Vol 258 (1) ◽

pp. 187-192 ◽

Cited By ~ 14

Author(s):

R Brandsch ◽

V Bichler ◽

B Krauss

Keyword(s):

Proteinase Inhibitors ◽

Cysteine Proteinase ◽

Cysteine Proteinase Inhibitor ◽

Western Blots ◽

E Coli ◽

Cell Extracts ◽

Arthrobacter Oxidans ◽

Cloned Gene

Expression of the 6-hydroxy-D-nicotine oxidase (6-HDNO) gene from Arthrobacter oxidans cloned into Escherichia coli showed a marked temperature-dependence. Transformed E. coli cells grown at 30 degrees C exhibited a several-fold higher 6-HDNO activity than did cells grown at 37 degrees C. This effect did not depend on the promoter used for expression of the cloned gene in E. coli, nor was it an effect of 6-HDNO mRNA instability at 37 degrees C. Studies performed in vivo and in vitro revealed that an increased susceptibility of apo-6-HDNO to proteolytic attack at 37 degrees C was responsible for the observed phenomenon. Extracts from cells grown at 37 degrees C showed on Western blots a decrease in immunologically detectable 6-HDNO polypeptide when compared with extracts from cells grown at 30 degrees C. The 6-HDNO polypeptide is covalently modified by attachment of the cofactor FAD to a histidine residue. It could be shown that covalent flavinylation of the apoenzyme in vitro, i.e. formation of holoenzyme, by incubation of cell extracts with FAD and phosphoenolpyruvate protected the 6-HDNO polypeptide from degradation at 37 degrees C. Of a variety of proteinase inhibitors tested only the cysteine-proteinase inhibitor L-3-trans-carboxyoxiran-2-carbonyl-L-leucylagmatine (E64) prevented degradation, by up to 70%, of the apoenzyme.

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Inhibition of lipopolysacharide induced osteoclast formation and bone resorption in vitro and in vivo in mice by cystatin C

Bone Abstracts ◽

10.1530/boneabs.1.pp225 ◽

2013 ◽

Author(s):

Stralberg Fredrik ◽

Catharina Lindholm ◽

Erik Lindstrom ◽

Franciszek Kasprzykowski ◽

Paul Saftig ◽

...

Keyword(s):

Bone Resorption ◽

Cystatin C ◽

Osteoclast Formation

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Osteoclast-mediated bone resorption is controlled by a compensatory network of secreted and membrane-tethered metalloproteinases

Science Translational Medicine ◽

10.1126/scitranslmed.aaw6143 ◽

2020 ◽

Vol 12 (529) ◽

pp. eaaw6143 ◽

Cited By ~ 11

Author(s):

Lingxin Zhu ◽

Yi Tang ◽

Xiao-Yan Li ◽

Evan T. Keller ◽

Jingwen Yang ◽

...

Keyword(s):

Bone Resorption ◽

Matrix Metalloproteinase ◽

Cysteine Proteinase ◽

Cathepsin K ◽

Collagenolytic Activity ◽

Type I ◽

Double Knockout ◽

Human Osteoclasts

Osteoclasts actively remodel both the mineral and proteinaceous components of bone during normal growth and development as well as pathologic states ranging from osteoporosis to bone metastasis. The cysteine proteinase cathepsin K confers osteoclasts with potent type I collagenolytic activity; however, cathepsin K–null mice, as well as cathepsin K–mutant humans, continue to remodel bone and degrade collagen by as-yet-undefined effectors. Here, we identify a cathepsin K–independent collagenolytic system in osteoclasts that is composed of a functionally redundant network of the secreted matrix metalloproteinase MMP9 and the membrane-anchored matrix metalloproteinase MMP14. Unexpectedly, whereas deleting either of the proteinases individually leaves bone resorption intact, dual targeting of Mmp9 and Mmp14 inhibited the resorptive activity of mouse osteoclasts in vitro and in vivo and human osteoclasts in vitro. In vivo, Mmp9/Mmp14 conditional double-knockout mice exhibited marked increases in bone density and displayed a highly protected status against either parathyroid hormone– or ovariectomy-induced pathologic bone loss. Together, these studies characterize a collagenolytic system operative in mouse and human osteoclasts and identify the MMP9/MMP14 axis as a potential target for therapeutic interventions for bone-wasting disease states.

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Activation of procathepsin B in human hepatoma cells: the conversion into the mature enzyme relies on the action of cathepsin B itself

Biochemical Journal ◽

10.1042/bj2930437 ◽

1993 ◽

Vol 293 (2) ◽

pp. 437-442 ◽

Cited By ~ 51

Author(s):

L Mach ◽

H Schwihla ◽

K Stüwe ◽

A D Rowan ◽

J S Mort ◽

...

Keyword(s):

Cathepsin B ◽

Mammalian Cells ◽

Proteinase Inhibitors ◽

Cysteine Proteinase ◽

Human Hepatoma ◽

Single Chain ◽

Human Hepatoma Cells ◽

Chain Form

In order to elucidate the processing mechanism of the lysosomal cysteine proteinase, cathepsin B, in mammalian cells, recombinant rat and human cathepsin B precursors were expressed in Saccharomyces cerevisiae. The active-site cysteine residue was changed to serine to prevent autoprocessing. When the purified proenzymes were incubated with the soluble fraction of postnuclear organelles obtained from human hepatoma HepG2 cells, processing to a 33 kDa form corresponding to the mature endogenous single-chain enzyme was observed. Inhibitors of metallo-, serine and aspartic proteinases exerted no significant effect on procathepsin B processing in vitro. However, the processing activity was effectively blocked by cysteine proteinase inhibitors, in particular E-64 and its cathepsin-B-selective derivative CA-074. Processing positions were identified by using anti-peptide antibodies specific for epitopes in the N- and C-terminal cleavage regions. The single-chain form produced in vitro was thus shown to contain an N-terminal extension of at least four residues relative to the mature lysosomal enzyme, as well as a C-terminal extension present in the proenzyme but usually absent in fully processed cathepsin B. On expression of the wild-type proenzyme in yeast, procathepsin B undergoes autoprocessing, yielding a single-chain form of the active enzyme, which contains similar N- and C-terminal extensions. These results indicate that maturation of procathepsin B in vivo in mammalian tissues relies on the proteolytic activity of cathepsin B itself.

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Influence of partial unfolding and aggregation of human stefin B (cystatin B) EPM1 mutants G50E and Q71P on selective cleavages by cathepsins B and S

Biological Chemistry ◽

10.1515/hsz-2012-0278 ◽

2013 ◽

Vol 394 (6) ◽

pp. 783-790 ◽

Cited By ~ 4

Author(s):

Mira Polajnar ◽

Robert Vidmar ◽

Matej Vizovišek ◽

Marko Fonović ◽

Nataša Kopitar-Jerala ◽

...

Keyword(s):

Proteinase Inhibitors ◽

Cysteine Proteinase ◽

Full Length ◽

Wild Type ◽

Cystatin B ◽

Cysteine Proteinase Inhibitors ◽

Sds Page ◽

Partial Unfolding ◽

Partially Unfolded

Abstract Human stefins and cystatins are physiologically important cysteine proteinase inhibitors, acting as a first line of defense against undesirable proteolysis. Mutations in the cystatin B gene cause a rare form of epilepsy EPM1. Its two missense mutants, G50E and Q71P, lack the inhibitory activity and are partially unfolded, which leads to changes in their aggregation behavior, both in vitro and in the cell. SDS-PAGE and MALDI-TOF mass spectrometry were used to follow the hydrolysis of human stefin B wild type, G50E and Q71P, by cathepsins B and S in vitro. Cathepsin S was found to degrade both mutants, with Q71P being degraded faster. This correlates with the openness of the protein structure, Q71P having more exposed hydrophobic surfaces. Cathepsin B acted more selectively, degrading G50E into smaller fragments, while still leaving a portion of the full-length protein intact. Q71P was cleaved only at the exposed N-terminal end. The co-localization of stefin B wild type and EPM1 mutants with cathepsins showed that cathepsins accumulate around the aggregates formed by the EPM1 mutants. We hypothesize that the aggregation of both full-length mutants prevents the cathepsin molecule from accessing the substrate protein’s core, whereas the cleaved fragments would be expected to aggregate stronger.

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In vitro embryotoxicity of the cysteine proteinase inhibitors benzyloxycarbonyl-phenylalanine-alanine-diazomethane (Z-Phe-Ala-CHN2) and benzyloxycarbony1-phenylalanine-phenylalanine-diazomethane (Z-Phe-Phe-CHN2)

Teratology ◽

10.1002/tera.1420500307 ◽

1994 ◽

Vol 50 (3) ◽

pp. 214-228 ◽

Cited By ~ 27

Author(s):

Jeffrey L. Ambroso ◽

Craig Harris

Keyword(s):

Proteinase Inhibitors ◽

Cysteine Proteinase ◽

Cysteine Proteinase Inhibitors

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LY900009 Regulates RANKL-Induced Osteoclast Formation and LPS-Induced Bone Resorption

10.21203/rs.3.rs-132552/v1 ◽

2020 ◽

Author(s):

Tao Huang ◽

Congyun Zhao ◽

Yi Zhao ◽

Yuan Zhou ◽

Lei Wang ◽

...

Keyword(s):

Bone Resorption ◽

Suppressive Effect ◽

Inhibitory Effect ◽

Osteoclast Formation ◽

Dependent Manner ◽

In Vivo Experiments ◽

Secretase Inhibitor ◽

Formation Number

Abstract To investigate the suppressive function of LY900009, a potent-secretase inhibitor, on RANKL-induced osteoclastogenesis. The cytotoxicity of LY900009 was evaluated. The suppressive effect and possible molecular mechanism of LY900009 on RANKL-induced osteoclastogenesis was evaluated both in vitro and in vivo. The IC50 of LY900009 was 2.93 mM. LY900009 treatment at different doses (100 nM, 200 nM, and 400 nM) effectively reduced osteoclast formation (number and arear) in a dose-dependent manner. The qPCR result shows that LY900009 attenuates RANKL-induced osteoclast formation and NFATc1 protein expression. The in vivo experiments demonstrated the inhibitory effect of LY900009 on LPS-induced bone resorption. LY900009 could potently inhibit osteoclastogenesis and bone resorption by down-regulating Notch/MAPK/Akt - mediated NFATc1 reduction in vitro. In accordance with the in vitro observations, we confirmed that LY900009 attenuated LPS-induced osteolysis in mice. In conclusion, our findings indicate that Notch was a potential therapeutic target which could be used for osteolytic diseases treatment.

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