Showing papers on "Tartrate-resistant acid phosphatase published in 1998"
Journal Article•
TL;DR: The resorbing cells in RA exhibit a definitive osteoclastic phenotype, suggesting that pharmacological agents that inhibit osteoclast recruitment or activity are rational targets for blocking focal bone erosion in patients with RA and JRA.
Abstract: Focal resorption of bone at the bone-pannus interface is common in rheumatoid arthritis (RA) and juvenile rheumatoid arthritis (JRA) and can result in significant morbidity. However, the specific cellular and hormonal mechanisms involved in this process are not well established. We examined tissue sections from areas of bone erosion in patients with RA and JRA. Multinucleated cells (MNCs) were present in resorption lacunae in areas of calcified cartilage and in subchondral bone immediately adjacent to calcified cartilage, as previously described. mRNA for the calcitonin receptor (CTR) was localized to these MNCs in bone resorption lacunae, a finding that definitively identifies these cells as osteoclasts. These MNCs were also positive for tartrate-resistant acid phosphatase (TRAP) mRNA and TRAP enzymatic activity. Occasional mononuclear cells on the bone surface were also CTR positive. Mononuclear cells and MNCs not on bone surfaces were CTR negative. The restriction of CTR-positive cells to the surface of mineralized tissues suggests that bone and/or calcified cartilage provide signals that are critical for the differentiation of hematopoietic osteoclast precursors to fully differentiated osteoclasts. Some MNCs and mononuclear cells off bone and within invading tissues were TRAP positive. These cells likely represent the precursors of the CTR-TRAP-positive cells on bone. Parathyroid hormone receptor mRNA was present in cells with the phenotypic appearance of osteoblasts, in close proximity to MNCs, and in occasional cells within pannus tissue, but not in the MNCs in bone resorption lacunae. These findings demonstrate that osteoclasts within the rheumatoid lesion do not express parathyroid hormone receptor. In conclusion, the resorbing cells in RA exhibit a definitive osteoclastic phenotype, suggesting that pharmacological agents that inhibit osteoclast recruitment or activity are rational targets for blocking focal bone erosion in patients with RA and JRA.
508 citations
TL;DR: This study demonstrates that mouse spleen cells and monocytes form osteoclasts when cultured in the presence of macrophage-colony stimulating factor (M-CSF) and a soluble form of murine ODF (sODF), and shows that the same co-stimulation causes human osteoclast differentiation to occur.
Abstract: Both human and murine osteoclasts can be derived in vitro from hematopoietic cells or monocytes that are cocultured with osteoblasts or marrow-derived stromal cells. The osteoclastogenic stimulus provided by murine osteoblasts and marrow-derived stromal cells is now known to be mediated by osteoclast differentiation factor (ODF), a membrane-bound tumor necrosis factor-related ligand. This study demonstrates that mouse spleen cells and monocytes form osteoclasts when cultured in the presence of macrophage-colony stimulating factor (M-CSF) and a soluble form of murine ODF (sODF). Numerous multinucleated osteoclasts expressing tartrate resistant acid phosphatase (TRAP) and calcitonin receptor (CTR) formed within 7 days of culture and engaged in extensive lacunar bone resorption. Osteoclast number and bone resorption area was dependent on sODF concentration. Long-term cultured human monocytes also formed bone resorbing osteoclasts in response to co-stimulation by sODF and M-CSF, although this required more th...
431 citations
TL;DR: It is concluded that CA II is essential not only in bone resorption but also in osteoclast differentiation, in both processes, however, the crucial role ofCA II is at least partially due to the effect on the osteOClast pHi regulation.
131 citations
TL;DR: The findings in this study are the first to indicate a new biological activity of Gas6 and protein S as a direct regulator of osteoclastic function; they give an insight into the role of these vitamin K‐dependent ligands in bone resorption in vivo.
Abstract: Bone is continuously being formed and resorbed. This process is accomplished by the precise coordination of two cell types: osteoblasts and osteoclasts. Osteoclasts are large, multinucleated cells that are derived from the same hematopoietic precursors as macrophages. However, these bone-resorbing cells are difficult to study directly because of their relative inaccessibility. The purification of primary osteoclasts from rabbit bones by their adherent nature provides an opportunity for investigating the molecules in osteoclasts. We have examined the expression of receptor tyrosine kinase by polymerase chain reaction (PCR) and found that Tyro 3 was frequently identified from primary osteoclasts in PCR cloning. Immunohistochemistry revealed that Tyro 3 was expressed on the multinucleated osteoclasts which were positive for tartrate-resistant acid phosphatase (TRAP), but not on mononuclear TRAP-positive cells. The Tyro 3 ligand, Gas6, induced the phosphorylation of Tyro 3 receptors in osteoclasts in two to five min. Gas6 and protein S directly enhanced the bone resorbing activity of mature osteoclasts. This effect of Gas6 was inhibited by the addition of a tyrosine kinase inhibitor, herbimycin A. However, Gas6 did not affect the differentiation of osteoclasts from bone marrow cells. Gas6 and protein S are dependent on vitamin K, a cofactor for the enzyme responsible for carboxylation of glutamic acid residues. The findings in this study are the first to indicate a new biological activity of Gas6 and protein S as a direct regulator of osteoclastic function; they give an insight into the role of these vitamin K-dependent ligands in bone resorption in vivo.
88 citations
Journal Article•
TL;DR: Findings suggest the direct participation of osteoclast-like cells in the joint destruction of CIA, the locally enhanced activity of osteoporosis-like cell differentiation in arthritic paws, and the participation of prostaglandins and prostag landin-independent IL-6 in this differentiation.
Abstract: Objective To investigate the participation of osteoclast-like bone resorbing cells in the joint destruction of murine collagen induced arthritis (CIA). Methods After induction of CIA in DBA/1J mice, a histological time course study was conducted on paw sections stained for tartrate resistant acid phosphatase (TRAP), a marker of osteoclasts. Cells from arthritic paws were cultured in vitro with or without indomethacin (IM) or anti-interleukin 6 neutralizing antibody (anti-IL-6), and stained for TRAP. Levels of prostaglandin E2 (PGE2), IL-1beta, IL-6, and tumor necrosis factor-alpha in the culture supernatants were determined by ELISA. The bone resorbing ability of these cells was examined on dentine slices. In control experiments, cells of normal paws or of arthritic tibiae were cultured in the same manner. Results TRAP positive osteoclast-like cells were detected late in the development of bone lesions at every eroded front in the pannus-bone and the pannus-subchondral bone junctions of arthritic joints. In vitro, cells of arthritic paws formed bone resorbing osteoclast-like cells spontaneously. However, the control culture failed to form these cells. PGE2 and IL-6 were detected at higher levels in arthritic culture than in control culture. Although both indomethacin and anti-IL-6 reduced osteoclast-like cell formation and indomethacin inhibited PGE2 synthesis, indomethacin failed to reduce IL-6. Conclusion These findings suggest the direct participation of osteoclast-like cells in the joint destruction of CIA, the locally enhanced activity of osteoclast-like cell differentiation in arthritic paws, and the participation of prostaglandins and prostaglandin-independent IL-6 in this differentiation.
86 citations
TL;DR: The findings indicate that production of H2O2 by HD‐11EM is an important local factor involved in differentiation of HD‐ 11EM into osteoclast‐like cells, and suggest that H 2O2 may play a role in native osteocline differentiation.
Abstract: Osteoclasts are the principal cells responsible for bone resorption (Chambers and Horton, 1984), and maintenance of skeletal mass is dependent on the formation and functional activation of these cells. Osteoclasts are generally believed to be derived from hematopoietic bone marrow progenitor cells, which have the capacity to differentiate and circulate in the blood as mononuclear osteoclast precursors (Fischman and Hay, 1962; Gothlin and Ericsson, 1973; Walker, 1973, 1975; Kahn and Simmons, 1975; Coccia et al., 1980). Maintaining skeletal mass is therefore dependent on the continuous recruitment of osteoclast precursors from the blood to bone surfaces. At or near bone surfaces, the mononuclear osteoclasts undergo further differentiation, and fuse to form the classic multinucleated osteoclast capable of resorbing bone. Although progress has been made in unraveling the basic cellular mechanisms that regulate the formation and activity of osteoclasts, our understanding of how they differentiate from hematopoietic bone marrow progenitors and become functionally active is incomplete (for review see Marks, 1983; Zaidi et al., 1993b; Roodman, 1995; Athanasou, 1996; Suda et al., 1997).
Advances in our understanding of osteoclast differentiation are based in part on the introduction of methods for the isolation and long-term culture of osteoclast bone marrow progenitor cells (Testa et al., 1981; Burger et al., 1982; Suda et al., 1997). However, osteoclast progenitor cells are not readily identifiable and are often a subpopulation of cells that may vary from preparation to preparation. In addition, most long-term culture systems suffer from the problem of being heterogeneous populations of cells, including osteoblasts and stromal cells, making it difficult to study the actual signaling pathways and responses of the osteoclast population to osteotropic hormones or local factors.
Another approach in the study of osteoclast differentiation has been to develop clonal cell lines from bone marrow progenitor cells, which have a high potential to differentiate into osteoclast-like multinucleate cells when cultured in the presence of osteotropic or neuroendocrine hormones (Gattei et al., 1992; Chambers et al., 1993; Shin et al., 1995; Hsia et al., 1995; Frediani et al., 1996; Hsia and Hauschka, unpublished data). However, none of these cell lines, despite showing osteoclast-like characteristics, have been shown to excavate resorption lacunae when incubated on slices of bone or dentin. Hsia and Hauschka (Hsia et al., 1995; Hsia and Hauschka, unpublished data) have recently developed a clonal cell line designated HD-11EM from the polyclonal v-myc-transformed myelomonocytic cell line, HD-11 (Beug et al., 1979). The HD-11EM clonal line shows a high potential to differentiate into osteoclast-like cells in response to the osteotropic hormone, 1α,25-dihydroxyvitamin D3 (1α,25-(OH)2D3). A significant percentage of the cells that form in response to 1α,25-(OH)2D3 are multinucleated, express tartrate-resistant acid phosphatase (TRAP), and excavate resorption pits when cultured on dentin slices in the presence of chicken osteoblasts (Hsia and Hauschka, unpublished data). The HD-11EM clonal line is therefore appropriate for studying the signaling pathways and responses of osteoclast precursors to osteotropic hormones or local factors that can influence their differentiation. Although the HD-11EM is an avian cell line, and the number of calcitonin receptors that are present on mature avian osteoclasts is known to be significantly less than those present on rat osteoclasts, we feel this is an appropriate line to study osteoclast differentiation, given that avian and mammalian osteoclasts demonstrate more similarities than differences in their responses to various hormones and in their resorptive activity (for review see Osdoby et al., 1992).
The differentiation and activity of osteoclasts are influenced by a number of osteotropic hormones, cytokines, and local factors (for review see: Zaidi et al., 1993b; Roodman, 1995; Suda et al., 1995, 1997; Manolagas, 1995; Athanasou, 1996). Among the osteotropic hormones, 1α,25-(OH)2D3 acts mainly by enhancing the differentiation of committed osteoclast precursors, and the addition of 1α,25-(OH)2D3 is required for the formation of osteoclasts in in vitro long-term cultures containing osteoblasts or stromal cells (Raisz et al., 1972; Bar-Shavit et al., 1983; Roodman et al., 1985; Takahashi et al., 1988; Udagawa et al., 1990; Quinn et al., 1994). In addition to the more accepted role of an indirect action of 1α,25-(OH)2D3 on the expression of local factors by osteoblasts or stromal cells, studies with HD-11EM cells indicate that 1α,25-(OH)2D3 can act directly on osteoclast precursors to stimulate osteoclast formation (Hsia et al., 1995; Hsia and Hauschka, unpublished data).
One local factor that may be involved in regulation of osteoclast differentiation is the production of reactive oxygen species (ROS), including superoxide and H2O2, originating from endothelial cells, stromal cells, and osteoclasts themselves. Garrett et al. (1990) initially reported ROS production by mature osteoclasts, as measured by the reduction of nitroblue tetrazolium (NBT), in mouse calvarial cultures in response to exogenous parathyroid hormone, interleukin-1, tumor necrosis factor, and 1α,25-(OH)2D3. These investigators also correlated production of ROS with osteoclast formation and activation, noting that addition of superoxide dismutase (SOD), which depletes superoxide, blocked NBT reduction and formation of additional osteoclasts. Catalase, which removes H2O2, had no effect in their studies. In contrast, Suda et al. (Suda, 1991; Suda et al., 1993) found that addition of catalase inhibited the number of osteoclast-like cells formed from progenitor cells in mouse calvarial organ cultures stimulated with 1α,25-(OH)2D3. Addition of exogenous H2O2 to their cultures overcame the effect of catalase, whereas addition of SOD, which would also increase the amount of H2O2, increased the number of osteoclast-like cells. The relative contributions of superoxide and/or H2O2 toward the differentiation of osteoclasts from progenitor cells remain unclear, as do the cellular origins of ROS and the regulating mechanisms of osteotropic hormones and local factors in stimulating ROS production.
The present study tests the hypothesis that production of a local factor(s) by HD-11EM cells in response to 1α,25-(OH)2D3 mediates the differentiation of HD-11EM into osteoclast-like cells. We were able to establish that one of the early local factors produced by HD-11EM cells in response to 1α,25-(OH)2D3 was H2O2, and that treatment of the HD-11EM cells with glucose/glucose oxidase, a pure H2O2 generation system, alone stimulated an increase in TRAP mRNA expression, TRAP activity/cell, and multinucleated cell formation 24–48 hr after treatment. These results implicate H2O2 as being integrally involved in the 1α,25-(OH)2D3-mediated differentiation of HD-11EM into osteoclast-like cells.
85 citations
TL;DR: Results support the theory of using a minimal concentration of hydrogen peroxide as a chemical adjuvant in the surgical treatment of giant cell tumors of bone.
Abstract: This study investigates the efficacy of using hydrogen peroxide as adjuvant therapy after extended local curettage for benign giant cell tumors of bone. Hydrogen peroxide is used clinically as a chemical adjuvant for removal of residual tumor cells, presumably by effervescent cleansing with minimal damage to surrounding soft tissue and bone cells. This investigation examined the effects of hydrogen peroxide on giant cell tumor cells and osteoblasts grown in culture. Fresh fragments of histologically confirmed giant cell tumor tissue (six patients) and trabecular bone (one patient) were excised. Cells obtained from the fragments were grown in culture. Confluent cell cultures were exposed to saline (control) or hydrogen peroxide (0.1-1000 mm) for 2 minutes, and incubation was continued for 12, 24, or 48 hours without hydrogen peroxide. Protein content, deoxyribonucleic acid content, tartrate resistant acid phosphatase activity, and alkaline phosphatase activity were measured in the cell layers. The medium from the final 12 hours of each incubation period was used to evaluate lactate production. Cell lysis or death occurred after exposing giant cell tumor cells and osteoblasts to 100 mm and 30 mm hydrogen peroxide, respectively, concentrations substantially lower than the 3% (880 mm) hydrogen peroxide commonly used clinically. These results support the theory of using a minimal concentration of hydrogen peroxide as a chemical adjuvant in the surgical treatment of giant cell tumors of bone.
81 citations
TL;DR: Findings suggest that bone destruction associated with local infection by S. aureus is due to the stimulation of osteoclast formation induced by the action of the easily solubilized SAM, and could explain the large numbers of osteoclasts found in infarcted bone in osteomyelitis.
Abstract: SUMMARY Objective. Staphylococcus aureus is the cause of bone destruction in osteomyelitis, bacterial arthritis and orthopaedic implant failure. We have previously shown that gentle saline extraction of S. aureus has revealed the presence of an extremely potent stimulator of osteoclast activation in both the murine calvarial bone resorption assay and the isolated chick osteoclast resorption assay. In order to investigate the mechanism of action of this surface-associated material (SAM ), we have investigated its capacity to recruit osteoclasts. Methods. The murine bone marrow osteoclast recruitment assay was used. The ability of the recruited cells to resorb dentine slices was also investigated. Results. The SAM from S. aureus dose dependently stimulated tartrate-resistant acid phosphatase (TRAP)-positive osteoclast formation and pit formation on dentine slices. Neutralization of the cytokines tumour necrosis factor alpha and interleukin (IL)-6 totally inhibited, but antagonism of IL-1 only partially blocked, the stimulated maturation of osteoclast-like cells. Conclusion. These findings suggest that bone destruction associated with local infection by S. aureus is due to the stimulation of osteoclast formation induced by the action of the easily solubilized SAM, and could explain the large numbers of osteoclasts found in infarcted bone in osteomyelitis.
73 citations
TL;DR: The utility of a mouse model to study the molecular and cellular bases for the regeneration and remodeling of tissue is demonstrated and the surgical technique and frame design are described.
73 citations
TL;DR: It is concluded that TRAP circulates in the serum as part of a complex, which also contains Ca2+, and that TR AP‐immunoassay is a potentially useful method for determining bone resorption rates, as long as the complex is destroyed before the assay.
Abstract: Osteoclasts secrete tartrate-resistant acid phosphatase (TRAP) to the circulation, where the amount of TRAP is expected to correlate with the bone resorption rate. We have developed two monoclonal antibodies, O1A and J1B, using purified human bone TRAP as antigen. The antibodies recognized different epitopes, allowing us to develop a two-site fluoroimmunoassay. The immunoreactivity in fresh serum specimens was less than 10% of the concentrations measured from the same specimens after 24 h of storage at 4 degrees C, or after addition of 5 mM EDTA or EGTA to them. When fresh serum was gel filtrated using Sephacryl S-200 column, all of the enzyme eluted in the void volume as a complex with a molecular weight of more than 250 kDa. If the serum was treated with EDTA before the gel filtration, the complex was destroyed and the enzyme eluted in fractions corresponding to a molecular weight of 30 kDa, the size of monomeric purified human bone TRAP. The immunoassay was used to measure TRAP concentrations from serum samples that had been stored at 4 degrees C for 24 h. According to the assay, premenopausal women had 13.1 +/- 3.1, postmenopausal women 17.6 +/- 4.2, and children 32.6 +/- 12.2 microg TRAP/l of serum. We conclude that TRAP circulates in the serum as part of a complex, which also contains Ca2+, and that TRAP-immunoassay is a potentially useful method for determining bone resorption rates, as long as the complex is destroyed before the assay.
72 citations
TL;DR: In this in vivo model, it took only 6 weeks for proliferative fibrous tissue to form, which penetrated the subchondral bone to communicate with the bone marrow and suggests that tumor necrosis factor alpha may play a major role in Fibrous tissue formation and osteoclastic bone resorption caused by wear debris around implants.
Abstract: Several in vivo studies have investigated bone to implant interface tissues after a single injection of wear particles into animal joints. However, in the clinical setting, wear debris are generated continuously rather than periodically. Histomorphologic findings and the gene expression of bone resorbing cytokines in reactive membranes were analyzed in response to continuous infusion of polyethylene particles with an osmotic pump. In this model, it took only 6 weeks for proliferative fibrous tissue to form, which penetrated the subchondral bone to communicate with the bone marrow. In addition, tartrate resistant acid phosphatase positive osteoclastlike cells were found in empty lacunae. According to the analysis of gene expression, only tumor necrosis factor alpha messenger ribonucleic acid was expressed specifically in the fibrous tissue of rats infused with particles from 4 weeks after surgery. This finding suggests that tumor necrosis factor alpha may play a major role in fibrous tissue formation and osteoclastic bone resorption caused by wear debris around implants. In addition, this in vivo model seems useful for investigating the mechanism of membrane formation and associated bone resorption around implants in a situation more similar to human prostheses than in conventional models.
TL;DR: Results strongly suggest that anti–FRP‐1 antibody‐induced multinucleated cells are osteoclasts, as FRP‐ 1 antigens were detected in osteoclast isolated from human bone and in the osteoclast‐like cells obtained from human giant cell tumors of bone.
Abstract: We have developed a new and simple system of human osteoclast formation by fusing peripheral blood monocytes with anti-Fusion Regulatory Protein-1 (anti-FRP-1) monoclonal antibody (mAb). When human blood monocytes were cultured in the presence of anti-FRP-1/CD98 mAbs, polykaryocytes began to appear at approximately 15 h and increased in size with time until 3-4 days of incubation with anti-FRP-1 mAb. These fused cells showed positive staining in tartrate-resistant acid phosphatase, possessed numerous calcitonin receptors, and were capable of bone resorption. These results strongly suggest that anti-FRP-1 antibody-induced multinucleated cells are osteoclasts. Furthermore, FRP-1 antigens were detected in osteoclasts isolated from human bone and in the osteoclast-like cells obtained from human giant cell tumors of bone.
TL;DR: The results indicate that chondroclasts and osteoclasts differ, not only with respect to location but possibly also by mode of action.
Abstract: The aim of the present study was to characterize cells involved in resorption during endochondral bone formation. We investigated whether the cells involved in cartilage breakdown at the epiphyseal/metaphyseal border, i.e., chondroclasts, share the characteristics of bone/cartilage-resorbing osteoclasts at the metaphyseal/diaphyseal border regarding ultrastructural features and functional activity. Morphometric evaluation showed that chondroclasts do not form ruffled borders and clear zones, i.e., well-known resorption characteristics, to the same extent as osteoclasts, present at the lower metaphysis. Instead, chondroclasts tend to express an undifferentiated surface adjacent to the matrix, not structurally different from the basolateral plasma membrane. Tartrate-resistant acid phosphatase (TRAP) was used as a marker for functional activity. Immunohistochemical staining by light microscopy was strong in both chondroclasts and in osteoclasts. Furthermore, in situ hybridization revealed large amounts of TRAP mRNA in chondroclasts as well as in osteoclasts. Ultrastructural immunohistochemistry suggests extensive secretion of the TRAP enzyme in the ruffled border area of both chondroclasts and osteoclasts. Intracellular accumulation was seen particularly in chondroclasts, possibly as a consequence of a relative disinclination to develop a ruffled border. Thus, semiquantitative estimation of TRAP distribution showed an inverse relationship between extracellular and intracellular TRAP in chondroclasts and osteoclasts. These results indicate that chondroclasts and osteoclasts differ, not only with respect to location but possibly also by mode of action. The observed differences may reflect the maturation sequence of these multinucleated cells when associated with different metaphyseal trabecular surfaces.
TL;DR: The results indicate that the effects of GH stimulation on osteoclastic resorption in vitro are mediated via local IGF-I secretion by stromal cells such as osteoblasts.
TL;DR: Cathepsin K expression during embryogenesis occurred only following the onset of osteoclast differentiation, and in situ hybridization studies in adult human tissues demonstrated high and specific expression in osteoclasts.
Abstract: We recently identified a novel cysteine protease, cathepsin K, by random sequencing of an osteoclast cDNA library, and in situ hybridization studies in adult human tissues demonstrated high and specific expression in osteoclasts. To determine whether the expression of cathepsin K mRNA during mouse embryogenesis was more widespread, cryostat sections of early (day 11-13) and late (day 15-17) mouse fetuses were analyzed by in situ hybridization. Serial cross-sections were collected through each fetus, and co-reacted for tartrate-resistant acid phosphatase (TRAP) and nonspecific esterase (NSE), selective markers for the osteoclast, and precursor cells derived from the macrophage/monocyte lineage, respectively. In the 11-13 day fetuses, cathepsin K mRNA was not expressed in any extraskeletal tissue; at this stage of embryogenesis, no osteoclasts are present. However, in the 15-17 day fetuses, a distinctive, developmental stage-dependent pattern of cathepsin K expression was observed in osteoclasts and preosteoclasts at sites of cartilage and bone modeling. Cathepsin K positive osteoclasts differentiated within a peripheral zone of the osteogenic stacked cell layer of the cartilage rudiments (prior to ossification), migrated and/or resorbed the bone collar, and invaded the cartilage core. Furthermore, following the invasive penetration of vasculature into the degenerating cartilage core, the calcified cartilage was resorbed by cathepsin K positive mononuclear osteoclast precursors (NSE+ve, negligible TRAP); cells positive for both enzymes were identified indicative of osteoclast differentiation. The deposition of bone by osteoblasts onto the cartilage remnants is followed by mononucleated and multinucleated osteoclastic resorption; these osteoclasts demonstrated intense cathepsin K expression. Similar expression patterns were observed at sites of intramembranous ossification. No expression was observed in chondrocytes, osteoblasts, marrow, or in any other nonskeletal tissue at these time points. These data indicated that cathepsin K expression during embryogenesis occurred only following the onset of osteoclast differentiation.
TL;DR: The expression of IL-6R and gp130 in bone marrow cells during osteoclast differentiation and TRAP-positive multinucleated cells during tartrate-resistant acid phosphatase (TRAP) formation are investigated to suggest that gp130 and IL- 6R, as well asIL-6, are involved in the formation and activation of osteoclasts.
TL;DR: The results suggest that redox-regulated PTPase activity may be a major function of TRAP in vivo and suggest that the newly synthesized enzyme is purple and reduced in the cell to the functionally active pink form.
TL;DR: The mRNA levels of estrogen receptor a, TRACP and CA II genes in neonatal rat bone imprints are investigated and suggest that estrogen may have a direct role in modulating the recruitment of osteoclast precursor cells during osteoclineogenesis.
Abstract: Increasing evidence suggests that estrogen deficiency in women promotes the expansion of populations of bone marrow cells that differentiate into osteoclasts under the influence of osteotropic hormones and local factors. A progressive cytoplasmic accumulation of osteoclastic bone resorbing enzymes, such as tartrate-resistant acid phosphatase (TRACP) and carbonic anhydrase II (CA II), characterizes osteoclast differentiation. To evaluate the possibility that estrogen may have a direct effect on osteoclast precursor cells, we investigated the mRNA levels of estrogen receptor a (ERa), TRACP and CA II genes in neonatal rat bone imprints by fluorescence in situ hybridization and confocal microscopy. Morphological assessment of bone imprints has shown that the putative mononuclear osteoclast precursor cells (MOPC) display strongly basophilic cytoplasm and a low nuclear/cytoplasmic ratio, while some of these cells possess pale-staining ruffled border regions similar to those observed in osteoclasts. Both CA II and TRACP mRNAs were detected in putative MOPC as well as multinuclear osteoclasts. The gene transcripts were mainly located in the cytoplasm of these cells. To determine whether these putative MOPC possess ER mRNA, a 637 base pair antisense ER riboprobe was used. The results indicated that MOPC which show TRACP reactivity express high levels of ER gene transcripts in their cytoplasm. In contrast, only a few multinuclear osteoclasts in the bone imprints possessed ER gene transcripts. Interestingly, the levels of ER mRNA in these multinuclear osteoclasts were very low compared with those in the putative MOPC. Treatment with RNase prior to hybridization resulted in a significant loss of signal in these cells. The results of these studies suggest that estrogen may have a direct role in modulating the recruitment of osteoclast precursor cells during osteoclastogenesis.
TL;DR: The structure of human prostatic acid phosphatase, noncovalently bound inN-propyl-l-tartramate, is used to develop inhibitors with higher specificity and potency thanl(+)-tartrate.
TL;DR: Both mRNA and protein for PTHrP were expressed in osteoclasts in sections of bone or joints from patients with Paget's disease, rheumatoid arthritis, and osteoarthritis, raising the possibility that PTHRP might participate in osteoclast function.
TL;DR: It is found that interleukin (IL)-13 as well as IL-4 induced peripheral blood monocytes and monoblastic cell line, UG3, to differentiate into MGCs in the presence of macrophage colony-stimulating factor (M-CSF), while IL-2, IL-7 or IL-10 did not.
TL;DR: The activity of tartrate-resistant and fluoride-sensitive acid phosphatase (TrFsACP) was measured by using 2,6-dichloro-4-acetylphenyl phosphate as substrate at pH 6.2 and the highest mean TrFsACP activity was found among children younger than 15 years, followed by that in elderly subjects (older than 60).
Abstract: We describe an improved method for the kinetic measurement of tartrate-resistant acid phosphatase (TrACP; EC [3.1.3.2][1]) activity in serum. Of the TrACP derived from erythrocytes, platelets, and macrophages (osteoclasts and others), that from the first two sources is also resistant to fluoride, whereas skeletal TrACP is sensitive to fluoride. Thus, osteoclast-derived TrACP can be measured specifically by exploiting its sensitivity to fluoride. We measured the activity of tartrate-resistant and fluoride-sensitive acid phosphatase (TrFsACP) by using 2,6-dichloro-4-acetylphenyl phosphate as substrate at pH 6.2. The activity of TrFsACP in serum was increased by adding hexadimethrine bromide (Polybrene) to the reaction mixture. This method was not influenced by hemolysis with hemoglobin concentrations as great as 0.9 g/L. The mean ± SD values of TrFsACP activity by this method were 20.4 ± 2.8 and 16.4 ± 2.3 U/L for young (ages 20–29 years) men (n = 34) and women (n = 50), respectively. The highest mean TrFsACP activity was found among children younger than 15 years, followed by that in elderly subjects (older than 60).
[1]: pending:yes
TL;DR: It is suggested that TRAP co-exists with TSAP, and is located in lysosomes and secretory granules in osteoblasts, and the similarity of the distribution patterns of TRAP and TSAP is similar.
Abstract: Ultrastructural localization of tartrate-resistant acid phosphatase (TRAP) activity in metaphyseal osteoblasts of young rats was detected and compared with the localization of tartrate-sensitive acid phosphatase (TSAP). TRAP activity was detected in lysosomes and Golgi complex, such as Golgi lamellae, vacuoles and vesicles. TSAP, detected as beta-glycerophosphatase, was also located in Golgi lamellae, vacuoles and vesicles, and in lysosomes. Based on the ultrastructural features of TRAP-positive structures and the similarity of the distribution patterns of TRAP and TSAP, it is suggested that TRAP co-exists with TSAP, and is located in lysosomes and secretory granules in osteoblasts.
TL;DR: The results suggest that, although the deterioration in the osteoblastic function can be commonly observed in NID DM animal models, the osteoclastic function is heterogeneous under NIDDM conditions.
Abstract: In order to investigate the pathogenic role of bone resorption by osteoclasts in altered bone metabolism in non–insulin-dependent diabetes mellitus (NIDDM), the circulating levels of tartrate resistant acid phosphatase (TRACP) were simultaneously determined with osteocalcin, in rat models of NIDDM, i.e., genetic Wistar fatty rats and neonatally streptozotocin-induced diabetic rats (NSZ rats). In Wistar fatty rats exhibiting hyperglycemia and hyperinsulinemia, plasma TRACP was 40.0 ± 0.4 U/l (mean ± SE), significantly higher than that of 32.8 ± 1.3 U/l in their lean littermates ( p p p p p These results suggest that, although the deterioration in the osteoblastic function can be commonly observed in NIDDM animal models, the osteoclastic function is heterogeneous under NIDDM conditions.
TL;DR: The authors investigated changes in the location and number of osteoclasts and their precursors during the development of articular lesions in type II collagen-induced arthritis in mice using tartrate-resistant acid phosphatase (TRAP) staining.
Abstract: The authors investigated changes in the location and number of osteoclasts and their precursors during the development of articular lesions in type II collagen-induced arthritis in mice using tartrate-resistant acid phosphatase (TRAP) staining. The limb joints were examined at 6 to 15 weeks after the second immunization. The number of TRAP-positive cells increased as the articular lesions progressed. TRAP-positive macrophage-like cells were found in the hyperplastic synovial tissue and bone marrow stroma in the early stage. In the advanced stage, in addition to many TRAP-positive osteoclasts on the bone surface, TRAP-positive macrophage-like cells were observed in the pannus apart from the bone surface in the pannus-joint junctions. The above mentioned TRAP-positive macrophage-like cells are considered to be osteoclast precursors.
TL;DR: A patient with adult type hypophosphatasia for over ten years who has also shown a persistently elevated tartrate resistant acid phosphatase activity (TRAP) without any obvious cause is followed, characterisation of this TRAP by polyacrylamide gel electrophoresis showed migration to band 5 position.
Journal Article•
TL;DR: In Canarian women, VDR genotype is not associated with changes in biochemical markers of bone remodelling or in bone mass or with the presence of osteoporosis or osteopOrotic fractures.
Abstract: BACKGROUND: Genetic factors condition an important part of bone mass. The role of vitamin D receptor polymorphism (VDR) as genetic marker of osteoporosis is a matter of discussion. We have studied the possible influence of VDR on bone remodelling, calciotropic hormones, on the presence of osteoporosis and osteoporotic bone fractures. PATIENTS, CONTROL POPULATION AND METHODS: A case-control study. We have studied a total of 127 postmenopausal Canarian women from Canary Islands, Spain; 66 healthy controls and 61 with the diagnosis of osteoporosis, which was made by clinical, radiological and densitometric criteria. 17 osteoporotic women have had a fracture: Colles, hip or vertebral (spinal deformity index) fracture. VDR were determined by PCR directed to demonstrate the presence (b) or absence (B) of a restriction target for Bsml in intron 7. We analyzed some biochemical markers of bone remodelling: serum levels of alkaline phosphatase, tartrate resistant acid phosphatase and urine ratios of calcium/creatinine and hydroxyproline/creatinine. We also determined calciotropic hormones: parathyroid hormone and calcitonin. Bone mass was measured by DEXA and TC. RESULTS: There were no significant differences in either biochemical bone remodelling markers or in bone mass between the three genotypes: bb, Bb and BB, either in controls or in osteoporotic women with the exception of alkaline phosphatase which had a significative increase compared to control in women with unfavorable alleles distribution (bB and BB). Distribution of genotypes was similar between controls and osteoporotic women, with or without fractures. CONCLUSIONS: In Canarian women, VDR genotype is not associated with changes in biochemical markers of bone remodelling or in bone mass or with the presence of osteoporosis or osteoporotic fractures.
TL;DR: It is concluded that the coloration of the bone matrix is probably caused by diffusion of the dye and reaction product and is, therefore, artifactual.
Abstract: It has been suggested that acid phosphatase activity is present in newly formed bone matrix at sites of endochondral ossification in rabbit fracture calluses. Because acid phosphatases are usually found intracellularly, it was decided to test this possibility more rigorously. Tissue from 10- and 14-day healing rabbit fractures was subjected to a series of critical tests for acid phosphatases with a pH optimum of 5.0. Fluoride, tartrate and molybdate were used as potential inhibitors of acid phosphatase activity. The effects of several counterstaining protocols were also investigated. A fluoride- and tartrate-resistant acid phosphatase is located in osteoclasts and mononuclear phagocytes. Diffuse staining of the bone matrix is seen, but it is dependent upon the length of incubation in the substrate medium and the distance from the acid phosphatase-reacting cells. It is concluded that the coloration of the bone matrix is probably caused by diffusion of the dye and reaction product and is, therefore, artifactual. © 1998 Chapman & Hall
TL;DR: Two unique antibodies to human TRAP purified from hairy cell leukemia spleen are developed, suitable for immunohistochemistry of osteoclasts and macrophages and failed to immunoprecipitate mouse or rat TRAP activity.
Abstract: Tartrate-resistant acid phosphatase (TRAP) is expressed abundantly by osteoclasts and is required for bone resorption. This enzyme is emerging as an important biomarker in bone pathology, both for histochemical identification of osteoclasts and as a serum marker of osteoclast activity and increased bone turnover. Rat and mouse models are becoming popular systems for studying osteoclast development, bone physiology and morphogenesis, and bone diseases such as osteoporosis. We have developed two unique antibodies to human TRAP purified from hairy cell leukemia spleen. Both antibodies (9C5 and 14G6) are suitable for immunohisto-chemistry of osteoclasts and macrophages. Only one (14G6) is capable of immunoprecipitating active TRAP from human cell lysates. Antibody 9C5 reacts with a denatured epitope of TRAP while antibody 14G6 probably reacts with a native, conformational determinant. The high degree of homology among TRAPs of various species predicts that these antibodies should be suitable for work in exper...
TL;DR: The results indicate that physiological concentrations of zinc alter bone resorptive rates in vitro by a mechanism that is dependent on DNA replication.
Abstract: The addition of physiological concentrations of zinc (25-200 (microg/dL) to Dulbecco's Modified Eagle's Medium containing tibiae from 19-d chick embryos resulted in a concentration-dependent increase in tibial content of tartrate-resistant acid phosphatase (TRAP) and an increase in bone resorption, as measured by tibial calcium release. This increase in bone resorption was additive to the resorptive effect resulting from the addition of 10(-9)-10(-7) M parathyroid hormone (PTH), but was not additive to similar effects produced by the addition of 10(-9)-10(-7) M prostaglandin E2 (PGE2). An inhibitor of prostaglandin synthesis, flurbiprofen (10[-6] M), did not influence the effect of zinc on bone resorption. However, the addition of 2,6-pyridinedicarboxylic acid (10[-3] M, 2,6-PDCA), a chelator of zinc, did attenuate the effects of zinc, as did the addition of an inhibitor of DNA replication (hydroxyurea, 10[-3] M). Hydroxyurea also attenuated the bone resorptive response to PGE2, but had no influence on the effects of PTH. These results indicate that physiological concentrations of zinc alter bone resorptive rates in vitro by a mechanism that is dependent on DNA replication.