A new cytometric method for the immunophenotypic characterization of bone‐derived human osteoclasts

Benito, Gloria Elena; Sánchez, María Luz; Pino‐Montes, Javier del; Calvo, José F.; Menéndez, Pablo; García-Marcos, M. A.; Osdoby, Philip; Órfão, Alberto

doi:10.1002/cyto.10139

Cited by 7 publications

(7 citation statements)

References 19 publications

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“…22 The effects of Ti(IV) on the osteoclastic activity and differentia-tion were also evaluated by measuring the enzymatic activity of TRAP using ELF97 as a substrate after 10 days of incubation. [19][20][21] As expected, expression of TRAP activity was not detected in untreated cultured monocytes. Analysis of in vitro generated osteoclasts showed TRAP activity at very high levels for $43% of the total cells, at lower levels for the majority of cells.…”

Section: Facs Analysis For Trap Activity In Monocytes and Osteoclastssupporting

confidence: 75%

“…All cells maintained a high level of CD45 expression 22. The effects of Ti(IV) on the osteoclastic activity and differentiation were also evaluated by measuring the enzymatic activity of TRAP using ELF97 as a substrate after 10 days of incubation 19–21. As expected, expression of TRAP activity was not detected in untreated cultured monocytes.…”

Section: Resultsmentioning

confidence: 69%

“…The aim of this study was to test the hypothesis that Ti(IV) ions, which represent the majority and most stable form of titanium ions released by biocorrosion from titanium implants, induce osteoclast differentiation in blood monocytes to become osteo‐resorptive cells. For this purpose, TRAP expression and activity was assessed 19–21. In addition, the bone resorptive function of the cells was evaluated by using dentin slide cultures.…”

Section: Introductionmentioning

confidence: 99%

“…For this purpose, TRAP expression and activity was assessed. [19][20][21] In addition, the bone resorptive function of the cells was evaluated by using dentin slide cultures.…”

Section: Introductionmentioning

confidence: 99%

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Titanium IV ions induced human osteoclast differentiation and enhanced bone resorption in vitro

Cadosch

Chan

Gautschi

et al. 2008

J Biomedical Materials Res

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There is increasing evidence that titanium (Ti) ions are released from orthopedic implants, with concentrations in the range of 1 microM in tissue and blood, and may play a role in aseptic loosening of orthopedic implants. This study investigated whether Ti(IV) ions induce differentiation of monocytic osteoclast precursors into osteo-resorptive multinucleated cells and influence the activation and function of in vitro generated osteoclasts. Human monocytes and in vitro generated osteoclasts were exposed to 1 microM Ti(IV) ions for 10 days. Thereafter, osteoclast differentiation, activation, and function were evaluated. Transcription of specific osteoclastic genes was measured using quantitative reverse transcription polymerase chain reactions, which showed increased expression of tartrate-resistant acid phosphatase (TRAP) in approximately 20% of Ti(IV)-treated monocytes. Detection and quantification of intracellular TRAP activity using ELF97 as a fluorescent substrate revealed a significant increase of TRAP-positive cells in Ti(IV)-treated monocytes. Additionally, as demonstrated on dentin slide cultures, Ti(IV)-treated monocytes became functional bone resorbing cells, significantly increasing their osteo-resorptive activity to similar levels as osteoclasts in vitro. These results suggest that Ti(IV) ions released by biocorrosion from orthopedic implants induce differentiation of monocytes toward mature, functional osteoclasts, which may well contribute the pathomechanism of aseptic loosening.

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Section: Facs Analysis For Trap Activity In Monocytes and Osteoclastssupporting

confidence: 75%

Section: Resultsmentioning

confidence: 69%

Section: Introductionmentioning

confidence: 99%

“…For this purpose, TRAP expression and activity was assessed. [19][20][21] In addition, the bone resorptive function of the cells was evaluated by using dentin slide cultures.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Titanium IV ions induced human osteoclast differentiation and enhanced bone resorption in vitro

Cadosch

Chan

Gautschi

et al. 2008

J Biomedical Materials Res

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“…Single-cell suspensions were obtained from spleen and lymph node tissues, by mechanical separation according to well-established procedures. 23 Bcl-2 expression was explored in both the normal and malignant B-cell subpopulations, by combined immunofluorescence stainings directed against cell surface and intracytoplasmic proteins, 24 using the following four-color combinations of monoclonal antibodies (MoAbs) directly conjugated with fluorescein isothiocyanate (FITC), phycoerythrin (PE), PE-cyanine 5 (PECy5) and allophycocyanine (APC): bcl-2-FITC (clone 124, DAKO cytomation, Glostrup, Denmark), either CD34-PE (clone HPCA2, Becton Dickinson Biosciences (BDB), San José, CA, USA) -in BM samples -or CD23-PE (clone EBVCS-5, BDB) -in PB, spleen and lymph node samples, CD19-PECy5 (clone SJ25C1, Caltag Laboratories, San Francisco, CA, USA) and CD38-APC (clone HB7, BDB). Cell fixation and permeabilization prior to bcl-2 protein staining was performed using the Fix&Perm reagent (Caltag Laboratories) according to the methods that have been previously described in detail.…”

Section: Flow Cytometry Immunophenotypic Studiesmentioning

confidence: 99%

Quantitative analysis of bcl-2 expression in normal and leukemic human B-cell differentiation

et al. 2004

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Lack of apoptosis has been linked to prolonged survival of malignant B cells expressing bcl-2. The aim of the present study was to analyze the amount of bcl-2 protein expressed along normal human B-cell maturation and to establish the frequency of aberrant bcl-2 expression in B-cell malignancies. In normal bone marrow (n ¼ 11), bcl-2 expression obtained by quantitative multiparametric flow cytometry was highly variable: very low in both CD34 þ and CD34 À B-cell precursors, high in mature B-lymphocytes and very high in plasma cells. Bcl-2 expression of mature B-lymphocytes from peripheral blood (n ¼ 10), spleen (n ¼ 8) and lymph node (n ¼ 5) was significantly higher (Po0.02) in CD23 À as compared to CD23 þ B cells, independent of the type of tissue analyzed. Upon comparison with normal human B-cell maturation, bcl-2 expression in neoplastic B cells from 144 patients was found to be aberrant in 66% of the cases, usually corresponding to bcl-2 overexpression (63%). Follicular lymphoma (FL) carrying t(14;18) and MALT lymphoma were the only diagnostic groups constantly showing overexpression of bcl-2. Bcl-2 overexpression was also frequently found in precursor B-acute lymphoblastic leukemia (84%), typical (77%) and atypical (75%) B-cell chronic lymphocytic leukemia, prolymphocytic leukemia (two of three cases), mantle cell lymphoma (55%), but not in t(14;18) À FL, splenic marginal zone lymphoma, Burkitt lymphoma and multiple myeloma.

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Cytometric analysis of protein expression and apoptosis in human primary cells with a novel microfluidic chip‐based system

Chan¹,

Luedke

Valer

et al. 2003

Cytometry Pt A

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Background:Work with primary cells is inherently limited by source availability and life span in culture. Flow cytometry offers extensive analytical opportunities but generally requires high cell numbers for an experiment. Methods: We have developed assays on a microfluidic system, which allow flow cytometric analysis of apoptosis and protein expression with a minimum number of fluorescently stained primary cells. In this setup, the cells are moved by pressure-driven flow inside a network of microfluidic channels and are analyzed individually by twochannel fluorescence detection. For some assays the staining reactions can be performed on-chip and the analysis is done without further washing steps. Results:We have successfully applied the assays to evaluate (a) activation of E-selectin (CD62E) expression by

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A new cytometric method for the immunophenotypic characterization of bone‐derived human osteoclasts

Cited by 7 publications

References 19 publications

Titanium IV ions induced human osteoclast differentiation and enhanced bone resorption in vitro

Titanium IV ions induced human osteoclast differentiation and enhanced bone resorption in vitro

Quantitative analysis of bcl-2 expression in normal and leukemic human B-cell differentiation

Cytometric analysis of protein expression and apoptosis in human primary cells with a novel microfluidic chip‐based system

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