Calcitonin down-regulates immediate cell signals induced in human osteoclast-like cells by the bone sialoprotein-IIA fragment through a postintegrin receptor mechanism

R. Paniccia, T. Riccioni, B. M. Zani, P. Zigrino, K. Scotlandi, A. Teti

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Abstract

Calcitonin (CT) is a peptide hormone that interacts with the cAMP-and phospholipase C-associated CT receptor subtypes. We investigated whether CT modulates the interaction of human tumoral osteoclast-like (GCT23) cells with a protein of the bone matrix, bone sialoprotein-II (BSP-II). Single GCT23 cells loaded with the intracellular Ca2+ indicator fura-2 were treated with the maximal active dose (300 μg/ml) of the 18-mer Arg-Gly-Asp (RGD)- containing BSP-IIA fragment, and the cytosolic free Ca2+ concentration ([Ca2+](i)) Was measured by dual wavelength microfluorometry. BSP-IIA stimulated an elevation in [Ca2+](i) , consisting mainly of a peak, followed by a rapid return toward baseline. Pretreatment with CT induced a modest elevation of [Ca2+](i). However, CT significantly inhibited the response to BSP-IIA in a dose-dependent manner. Maximal inhibition (90% vs. untreated) was observed in the micromolar range. The intracellular mechanisms leading to this effect were investigated by pretreatment of GCT23 cells with the cAMP permeant analog, (Bu2)cAMP, and the protein kinase-C-activating agent, 12-O-tetradecanoylphorbol 13-acetate. Similar to CT, both agents inhibited the response to 300 μg/ml BSP-IIA. The effect induced by CT was specific, because an increase in the extracellular Ca2+ concentration, which is also known to inhibit bone resorption, failed to modify the ability of BSP-IIA to alter [Ca2+ in GCT23 cells. To investigate whether the CT- induced alteration of BSp-IIA-dependent cell signals was due to a modification in the synthesis of cell surface receptors (integrins) for the extracellular matrix macromolecules, 1-h CT-treated [35S]methionine metabolically labeled GCT23 cell lysates were immunoprecipitated with anti- α3-, -α(v)-, -β1-, and -β3-integrin subunit antibodies. Autoradiography demonstrated that 10-7-10-6 M CT did not alter new synthesis of the and the α3β1 receptors. Similarly, CT did not affect surface expression of these receptors, assessed by enzyme-linked immunosorbent assay. Finally, no alteration of the adhesion rate and spreading of GCT23 cells onto BSP-IIA-coated substrates was observed. This indicates that CT-induced down-regulation of immediate cell signals prompted by BSP-IIA in GCT23 cells is a postintegrin receptor event.

Original languageEnglish
Pages (from-to)1177-1186
Number of pages10
JournalEndocrinology
Volume136
Issue number3
Publication statusPublished - 1995

Fingerprint

Integrin-Binding Sialoprotein
Calcitonin
Osteoclasts
Down-Regulation
Integrins
Calcitonin Receptors
Cytophotometry
Bone Matrix
Peptide Hormones
Cell Surface Receptors
Type C Phospholipases
Tetradecanoylphorbol Acetate
Bone Resorption
Autoradiography
Methionine
Protein Kinase C
Extracellular Matrix

ASJC Scopus subject areas

  • Endocrinology
  • Endocrinology, Diabetes and Metabolism

Cite this

Calcitonin down-regulates immediate cell signals induced in human osteoclast-like cells by the bone sialoprotein-IIA fragment through a postintegrin receptor mechanism. / Paniccia, R.; Riccioni, T.; Zani, B. M.; Zigrino, P.; Scotlandi, K.; Teti, A.

In: Endocrinology, Vol. 136, No. 3, 1995, p. 1177-1186.

Research output: Contribution to journalArticle

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abstract = "Calcitonin (CT) is a peptide hormone that interacts with the cAMP-and phospholipase C-associated CT receptor subtypes. We investigated whether CT modulates the interaction of human tumoral osteoclast-like (GCT23) cells with a protein of the bone matrix, bone sialoprotein-II (BSP-II). Single GCT23 cells loaded with the intracellular Ca2+ indicator fura-2 were treated with the maximal active dose (300 μg/ml) of the 18-mer Arg-Gly-Asp (RGD)- containing BSP-IIA fragment, and the cytosolic free Ca2+ concentration ([Ca2+](i)) Was measured by dual wavelength microfluorometry. BSP-IIA stimulated an elevation in [Ca2+](i) , consisting mainly of a peak, followed by a rapid return toward baseline. Pretreatment with CT induced a modest elevation of [Ca2+](i). However, CT significantly inhibited the response to BSP-IIA in a dose-dependent manner. Maximal inhibition (90{\%} vs. untreated) was observed in the micromolar range. The intracellular mechanisms leading to this effect were investigated by pretreatment of GCT23 cells with the cAMP permeant analog, (Bu2)cAMP, and the protein kinase-C-activating agent, 12-O-tetradecanoylphorbol 13-acetate. Similar to CT, both agents inhibited the response to 300 μg/ml BSP-IIA. The effect induced by CT was specific, because an increase in the extracellular Ca2+ concentration, which is also known to inhibit bone resorption, failed to modify the ability of BSP-IIA to alter [Ca2+ in GCT23 cells. To investigate whether the CT- induced alteration of BSp-IIA-dependent cell signals was due to a modification in the synthesis of cell surface receptors (integrins) for the extracellular matrix macromolecules, 1-h CT-treated [35S]methionine metabolically labeled GCT23 cell lysates were immunoprecipitated with anti- α3-, -α(v)-, -β1-, and -β3-integrin subunit antibodies. Autoradiography demonstrated that 10-7-10-6 M CT did not alter new synthesis of the and the α3β1 receptors. Similarly, CT did not affect surface expression of these receptors, assessed by enzyme-linked immunosorbent assay. Finally, no alteration of the adhesion rate and spreading of GCT23 cells onto BSP-IIA-coated substrates was observed. This indicates that CT-induced down-regulation of immediate cell signals prompted by BSP-IIA in GCT23 cells is a postintegrin receptor event.",
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T1 - Calcitonin down-regulates immediate cell signals induced in human osteoclast-like cells by the bone sialoprotein-IIA fragment through a postintegrin receptor mechanism

AU - Paniccia, R.

AU - Riccioni, T.

AU - Zani, B. M.

AU - Zigrino, P.

AU - Scotlandi, K.

AU - Teti, A.

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