Dephosphorylation of osteopontin and bone sialoprotein by osteoclastic tartrate-resistant acid phosphatase. Modulation of osteoclast adhesion in vitro.

Abstract. 1,25-Dihydroxy-19-nor-vitamin D2 (19-norD2), a new analog of 1,25(OH)2D3, suppresses parathyroid hormone in renal failure patients and in uremic rats but has less calcemic activity than 1,25(OH)2D3. Although 19-norD2 has high affinity for the vitamin D receptor and similar pharmacokinetics to those of 1,25(OH)2D3, it has much less bone resorbing activity in vivo. The intrinsic activity of 19-norD2 on osteoclastogenesis and activation of bone resorption in mouse bone marrow cultures was examined to determine the mechanism involved. 19-norD2 and 1,25(OH)2D3 (10 nM) were equivalent in stimulating the formation and maintenance of large multinucleated, tartrate-resistant acid phosphatase-positive cells. However, the amount of bone resorbed by osteoclasts stimulated by 10 nM 19-norD2, as measured by pit-forming assays, was reduced 62% compared with 10 nM 1,25(OH)2D3-stimulated osteoclasts (P < 0.05). This difference could not be attributed to enhanced catabolism or to downregulated vitamin D receptor. The rate of degradation of 19-norD2 in cultures was approximately 20% greater than 1,25(OH)2D3, not enough to account for the different effects on bone resorption. The VDR levels were identical in cultures that were treated with 19-norD2 and 1,25(OH)2D3. In summary, 19-norD2 is less effective than 1,25(OH)2D3 in stimulating mouse marrow osteoclasts to resorb bone. The reason for this difference is not clear but seems to involve the late maturation and/or activation of osteoclasts as the number of pits produced by each tartrate-resistant acid phosphatase-positive cell is reduced under stimulation by 19-norD2 compared with 1,25(OH)2D3.

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Tartrate-resistant acid phosphatase from human osteoclastomas is translated as a single polypeptide

Biochemical Journal ◽

10.1042/bj2770631 ◽

1991 ◽

Vol 277 (3) ◽

pp. 631-634 ◽

Cited By ~ 25

Author(s):

A R Hayman ◽

A J Dryden ◽

T J Chambers ◽

M J Warburton

Keyword(s):

Acid Phosphatase ◽

Peptide Bond ◽

Northern Blotting ◽

Tartrate Resistant Acid Phosphatase ◽

Acid Phosphatases ◽

Post Translational Modification ◽

Subunit Protein ◽

Single Polypeptide ◽

Single Peptide

Tartrate-resistant acid phosphatases have been isolated from a number of sources. These enzymes consist of one subunit (Mr 30,000-40,000) or two dissimilar subunits (Mr 15,000-20,000). Previously we isolated the enzyme from human osteoclastomas, as a two-subunit protein. By Northern blotting and hybridization with radiolabelled oligonucleotides corresponding to the N-terminal sequences of the two subunits, we demonstrate here that the enzyme is transcribed as one mRNA which is translated in vitro to produce a single polypeptide of approx. Mr 33,000. Transcription as a single mRNA species is also the case in other tissues. These results suggest that the osteoclastoma enzyme undergoes post-translational modification in the form of cleavage of a single peptide bond to give a disulphide-bonded two-subunit protein.

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Functional Dissociation of the Basolateral Transcytotic Compartment from the Apical Phago-lysosomal Compartment in Human Osteoclasts

Journal of Histochemistry & Cytochemistry ◽

10.1369/jhc.4a6476.2005 ◽

2005 ◽

Vol 53 (5) ◽

pp. 665-670 ◽

Cited By ~ 11

Author(s):

James Meagher ◽

René Zellweger ◽

Luis Filgueira

Keyword(s):

Staphylococcus Aureus ◽

Dendritic Cells ◽

Acid Phosphatase ◽

In Vitro Study ◽

Infectious Agent ◽

Tartrate Resistant Acid Phosphatase ◽

Lysosomal Compartment ◽

Acidic Compartment ◽

Human Osteoclasts

Tartrate-resistant acid phosphatase (TRAP) is essential for elimination of Staphylococcus aureus, the main infectious agent responsible for osteomyelitis. This in vitro study investigated uptake and processing of fluorescence-labeled S. aureus by human osteoclasts and dendritic cells. The cells were stained for TRAP and the acidic compartment using a fluorescence-based protocol. In dendritic cells, TRAP and bacteria were colocalized. In osteoclasts, there was no colocalization of bacteria, TRAP, or the acidic compartment, indicating that there are three distinct vesicular compartments: the apical phago-lysosomal compartment, the basal secretory compartment, and the basolateral transcytotic compartment. Dissociation of the TRAP-containing transcytotic vesicles from the apical phago-lysosomal compartment may restrain osteoclasts from eliminating S. aureus.

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Osteoclast 121F antigen expression during osteoblast conditioned medium induction of osteoclast-like cells in vitro: Relationship to calcitonin responsiveness, tartrate resistant acid phosphatase levels, and bone resorptive activity

Journal of Bone and Mineral Research ◽

10.1002/jbmr.5650100109 ◽

2009 ◽

Vol 10 (1) ◽

pp. 45-58 ◽

Cited By ~ 15

Author(s):

Patricia Collin-Osdoby ◽

Merry Jo Oursler ◽

Linda Rothe ◽

David Webber ◽

Fred Anderson ◽

...

Keyword(s):

Acid Phosphatase ◽

Conditioned Medium ◽

Antigen Expression ◽

Tartrate Resistant Acid Phosphatase ◽

Resorptive Activity

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Mice lacking tartrate-resistant acid phosphatase (Acp 5) have disrupted endochondral ossification and mild osteopetrosis

Development ◽

10.1242/dev.122.10.3151 ◽

1996 ◽

Vol 122 (10) ◽

pp. 3151-3162 ◽

Cited By ~ 1

Author(s):

A.R. Hayman ◽

S.J. Jones ◽

A. Boyde ◽

D. Foster ◽

W.H. Colledge ◽

...

Keyword(s):

Acid Phosphatase ◽

Endochondral Ossification ◽

Bone Matrix ◽

Axial Skeleton ◽

Mutant Mice ◽

Intrinsic Defect ◽

Tartrate Resistant Acid Phosphatase ◽

Long Bones ◽

Electron Imaging

Mature osteoclasts specifically express the purple, band 5 isozyme (Acp 5) of tartrate-resistant acid phosphatase, a binuclear metalloenzyme that can generate reactive oxygen species. The function of Acp 5 was investigated by targeted disruption of the gene in mice. Animals homozygous for the null Acp 5 allele had progressive foreshortening and deformity of the long bones and axial skeleton but apparently normal tooth eruption and skull plate development, indicating a role for Acp 5 in endochondral ossification. Histomorphometry and mineralization density analysis of backscattered electron imaging revealed widened and disorganized epiphyseal growth plates with delayed mineralization of cartilage in 6- to 8-week-old mutant mice. The membrane bones of the skull showed increased density at all ages examined, indicating defective osteoclastic bone turnover. Increased mineralization density was observed in the long bones of older animals which showed modelling deformities at their extremities: heterozygotes and homozygous Acp 5 mutant mice had tissue that was more mineralized and occupied a greater proportion of the bone in all regions. Thus the findings reflect a mild osteopetrosis due to an intrinsic defect of osteoclastic modelling activity that was confirmed in the resorption pit assay in vitro. We conclude that this bifunctional metalloprotein of the osteoclast is required for normal mineralization of cartilage in developing bones; it also maintains integrity and turnover of the adult skeleton by a critical contribution to bone matrix resorption.

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Rapid Screening Method for Osteoclast Differentiation in Vitro That Measures Tartrate-resistant Acid Phosphatase 5b Activity Secreted into the Culture Medium

Clinical Chemistry ◽

10.1093/clinchem/46.11.1751 ◽

2000 ◽

Vol 46 (11) ◽

pp. 1751-1754 ◽

Cited By ~ 78

Author(s):

Sari L Alatalo ◽

Jussi M Halleen ◽

Teuvo A Hentunen ◽

Jukka Mönkkönen ◽

H Kalervo Väänänen

Keyword(s):

Acid Phosphatase ◽

Culture Medium ◽

Osteoclast Differentiation ◽

Therapeutic Agents ◽

Precursor Cells ◽

Osteoclast Precursor ◽

Tartrate Resistant Acid Phosphatase ◽

Dependent Manner ◽

Trap 5B

Abstract Background: Osteoclasts secrete tartrate-resistant acid phosphatase (TRAP; EC 3.1.3.2) 5b into the circulation. We studied the release of TRAP 5b from osteoclasts using a mouse in vitro osteoclast differentiation assay. Methods: We developed and characterized a polyclonal antiserum in rabbits, using purified human osteoclastic TRAP 5b as antigen. The antiserum was specific for TRAP in Western analysis of mouse osteoclast culture medium and was used to develop an immunoassay. We cultured mouse bone marrow-derived osteoclast precursor cells for 3–7 days with or without clodronate in the presence of vitamin D and analyzed the number of osteoclasts formed and the amount of TRAP 5b activity released into the culture medium. Results: TRAP 5b activity was not secreted from osteoclast precursor cells. Addition of clodronate-containing liposomes decreased in a dose-dependent manner the number of osteoclasts and TRAP 5b activity released in 6-day cultures. The amount of TRAP 5b activity in the medium detected by the immunoassay correlated significantly with the number of osteoclasts formed (r = 0.94; P <0.0001; n = 120). Conclusions: The TRAP 5b immunoassay can be used to replace the laborious and time-consuming microscopic counting of osteoclasts in the osteoclast differentiation assay and to test the effects of potential therapeutic agents on osteoclast differentiation, enabling fast screening of large amounts of potential therapeutic agents.

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The Antiosteoporosis Effects of Zhuanggu Guanjie Pill In Vitro and In Vivo

BioMed Research International ◽

10.1155/2018/9075318 ◽

2018 ◽

Vol 2018 ◽

pp. 1-11 ◽

Cited By ~ 1

Author(s):

Li-juan Chai ◽

Yue Zhang ◽

Pan-yang Zhang ◽

Ya-nan Bi ◽

Xiao-mei Yuan ◽

...

Keyword(s):

Acid Phosphatase ◽

Bone Resorption ◽

Phosphatase Activity ◽

Bone Strength ◽

Acid Phosphatase Activity ◽

Ovariectomized Rats ◽

Tartrate Resistant Acid Phosphatase ◽

Trabecular Thickness

We investigated the beneficial effects and underlying mechanisms of Zhuanggu Guanjie (ZGGJ) pill in osteoporosis in vitro and in vivo. Bone marrow macrophages from 4–6-week-old mice were cultured in the presence of macrophage colony-stimulating factor (15 ng/mL) and receptor activator of nuclear factor-κB ligand (30 ng/mL). Osteoclast differentiation was determined by quantification of tartrate-resistant acid phosphatase activity. Gelatin zymography was used to detect the activity of matrix metalloproteinases in osteoclasts. Ovariectomized rats were administered orally with estradiol valerate or ZGGJ for 8 weeks. Blood was collected to measure serum indices. Tibiae were harvested to carry out bone microcomputed tomography scanning, histomorphological analysis, and bone strength determination. ZGGJ inhibited tartrate-resistant acid phosphatase activity, matrix metalloproteinase 9 expression, and bone resorption in vitro. At doses of 0.55, 1.1, and 2.2 g/kg, ZGGJ exerted significant osteoprotective effects including inhibition of bone turnover markers and improved tibia bone strength in ovariectomized rats. Microcomputed tomographic analysis showed that ZGGJ improved the trabecular architecture with increased connectivity density and trabecular thickness and decreased trabecular spacing. These results revealed that ZGGJ prevents bone loss induced by ovariectomy in rats and that inhibition of bone resorption is involved in the bone-protective effects of ZGGJ.

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