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Calcif Tissue Int. 2019 Oct 25. doi: 10.1007/s00223-019-00618-w. [Epub ahead of print]

A Novel Sandwich ELISA for Tartrate-Resistant Acid Phosphatase 5a and 5b Protein Reveals that Both Isoforms are Secreted by Differentiating Osteoclasts and Correlate to the Type I Collagen Degradation Marker CTX-I In Vivo and In Vitro.

Author information

1
Division of Pathology, Department of Laboratory Medicine, Karolinska Institutet, Alfred Nobels Allé, 8, 141 52, Stockholm, Sweden.
2
Mabtech AB, Box 1233, 131 28, Nacka Strand, Sweden.
3
Division of Pathology, Department of Laboratory Medicine, Karolinska Institutet, Alfred Nobels Allé, 8, 141 52, Stockholm, Sweden. tuomas.nareoja@ki.se.
4
Division of Pathology, Department of Laboratory Medicine, Karolinska Institutet, Alfred Nobels Allé, 8, 141 52, Stockholm, Sweden. pernilla.lang@ki.se.

Abstract

Tartrate-resistant acid phosphatase type 5 (TRAP) exists as two isoforms, 5a and 5b. 5b is a marker of osteoclast number and 5a of chronic inflammation; however, its association with bone resorption is unknown. In this study, a double-TRAP 5a/5b sandwich ELISA measuring 5a and 5b protein in the same sample was developed. TRAP 5a and 5b protein levels were evaluated as osteoclast differentiation/activity markers in serum and in culture, and their correlation to the resorption marker CTX-I was examined. Serum TRAP 5a and 5b concentrations in healthy men were 4.4 ± 0.6 ng/ml and 1.3 ± 0.2 ng/ml, respectively, and they correlated moderately to each other suggesting that their secretion is coupled under healthy conditions. A correlation was also observed between serum TRAP 5a and 5b with CTX-I, suggesting that both TRAP isoforms associate with osteoclast number. During osteoclast differentiation on plastic/bone, predominantly 5b increased in media/lysate from M-CSF/RANKL-stimulated CD14+ PBMCs. However, substantial levels of 5a were detected at later stages suggesting that both isoforms are secreted from differentiating OCs. More TRAP 5b was released on bone indicating a connection to osteoclast resorptive activity, and a peak in TRAP 5b/5a-ratio coincided with rapid CTX-I release. At the end of the culture period of M-CSF + RANKL-stimulated CD14+ PBMCs, there was a correlation between the secretion of TRAP 5a and 5b proteins with CTX-I. The correlation of not only 5b but also 5a with collagen degradation, both in serum and osteoclast cultures indicates that a considerable proportion of the TRAP 5a originates from osteoclasts and may reflect a hitherto undisclosed regulatory mechanism during bone resorption and bone remodeling.

KEYWORDS:

Acp5; Biomarker; Bone resorption; ELISA; Osteoclast

PMID:
31654098
DOI:
10.1007/s00223-019-00618-w

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