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RANKL regulates Fas expression and Fas-mediated apoptosis in osteoclasts.
J Bone Miner Res. 2005 Jan; 20(1):107-16.JB

Abstract

Osteoclast apoptosis is an influential determinant of osteoclast bone-resorbing activity. RANKL, a critical factor for osteoclastogenesis, is also important in osteoclast survival. However, the mechanisms by which RANKL prevents osteoclast apoptosis remain largely unknown.

INTRODUCTION

Fas, a death receptor, mediates apoptosis in multiple types of cells including osteoclasts. Here we report that RANKL acts as a survival factor in osteoclasts by downregulating Fas-mediated apoptosis and Fas expression in mature osteoclasts.

MATERIALS AND METHODS

RAW264.7 and mouse bone marrow macrophage/monocyte progenitors and progenitor-derived osteoclasts, in the presence of various concentrations of RANKL, were used in this study. Western blotting, semiquantitative RT-PCR, flow cytometry, nuclear staining, and a fluorescent caspase-3 activity assay were used to assess the effect of RANKL on Fas expression and Fas-mediated apoptosis. The involvement of NF-kappaB in the regulation of Fas by RANKL was analyzed by luciferase assay and EMSA.

RESULTS

Mature osteoclasts generated in the presence of a high concentration of RANKL (3.33 nM) failed to respond to Fas-induced apoptosis. The lack of responsiveness in mature osteoclasts is caused by the low level of Fas expression, as detected by both semiquantitative PCR and Western blotting. Fas protein and mRNA expression are inhibited by RANKL in concentration-dependent manners. The downregulation of Fas expression by RANKL is not because of modulation of the stability of Fas protein or mRNA. The regulation of Fas expression by RANKL is biphasic. During the early stage of osteoclastogenesis (1 day) when Fas is expressed at a very low level, RANKL upregulates Fas promoter activity by 2.4 +/- 0.1-fold in a concentration-dependent manner and increases Fas mRNA and protein. This event correlates with regulation of the binding activity of NF-kappaB to the Fas promoter by RANKL, as detected by EMSA. In osteoclast precursors, the induction of Fas promoter activity by RANKL was dramatically reduced when NF-kappaB binding sites on the Fas promoter were mutated.

CONCLUSION

RANKL upregulates Fas expression in osteoclast progenitors through NF-kappaB, making osteoclasts targets of Fas-stimulated apoptosis. In differentiated mature osteoclasts, RANKL reduces the levels of Fas expression and Fas-mediated apoptosis, acting as a survival factor.

Authors+Show Affiliations

Department of Pathology, University of Alabama at Birmingham, Birmingham, Alabama 35294-0007, USA.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

15619676

Citation

Wu, Xiaojun, et al. "RANKL Regulates Fas Expression and Fas-mediated Apoptosis in Osteoclasts." Journal of Bone and Mineral Research : the Official Journal of the American Society for Bone and Mineral Research, vol. 20, no. 1, 2005, pp. 107-16.
Wu X, Pan G, McKenna MA, et al. RANKL regulates Fas expression and Fas-mediated apoptosis in osteoclasts. J Bone Miner Res. 2005;20(1):107-16.
Wu, X., Pan, G., McKenna, M. A., Zayzafoon, M., Xiong, W. C., & McDonald, J. M. (2005). RANKL regulates Fas expression and Fas-mediated apoptosis in osteoclasts. Journal of Bone and Mineral Research : the Official Journal of the American Society for Bone and Mineral Research, 20(1), 107-16.
Wu X, et al. RANKL Regulates Fas Expression and Fas-mediated Apoptosis in Osteoclasts. J Bone Miner Res. 2005;20(1):107-16. PubMed PMID: 15619676.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - RANKL regulates Fas expression and Fas-mediated apoptosis in osteoclasts. AU - Wu,Xiaojun, AU - Pan,George, AU - McKenna,Margaret A, AU - Zayzafoon,Majd, AU - Xiong,Wen-Cheng, AU - McDonald,Jay M, Y1 - 2004/10/25/ PY - 2004/03/31/received PY - 2004/07/06/revised PY - 2004/08/12/accepted PY - 2004/12/28/pubmed PY - 2005/5/26/medline PY - 2004/12/28/entrez SP - 107 EP - 16 JF - Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research JO - J Bone Miner Res VL - 20 IS - 1 N2 - UNLABELLED: Osteoclast apoptosis is an influential determinant of osteoclast bone-resorbing activity. RANKL, a critical factor for osteoclastogenesis, is also important in osteoclast survival. However, the mechanisms by which RANKL prevents osteoclast apoptosis remain largely unknown. INTRODUCTION: Fas, a death receptor, mediates apoptosis in multiple types of cells including osteoclasts. Here we report that RANKL acts as a survival factor in osteoclasts by downregulating Fas-mediated apoptosis and Fas expression in mature osteoclasts. MATERIALS AND METHODS: RAW264.7 and mouse bone marrow macrophage/monocyte progenitors and progenitor-derived osteoclasts, in the presence of various concentrations of RANKL, were used in this study. Western blotting, semiquantitative RT-PCR, flow cytometry, nuclear staining, and a fluorescent caspase-3 activity assay were used to assess the effect of RANKL on Fas expression and Fas-mediated apoptosis. The involvement of NF-kappaB in the regulation of Fas by RANKL was analyzed by luciferase assay and EMSA. RESULTS: Mature osteoclasts generated in the presence of a high concentration of RANKL (3.33 nM) failed to respond to Fas-induced apoptosis. The lack of responsiveness in mature osteoclasts is caused by the low level of Fas expression, as detected by both semiquantitative PCR and Western blotting. Fas protein and mRNA expression are inhibited by RANKL in concentration-dependent manners. The downregulation of Fas expression by RANKL is not because of modulation of the stability of Fas protein or mRNA. The regulation of Fas expression by RANKL is biphasic. During the early stage of osteoclastogenesis (1 day) when Fas is expressed at a very low level, RANKL upregulates Fas promoter activity by 2.4 +/- 0.1-fold in a concentration-dependent manner and increases Fas mRNA and protein. This event correlates with regulation of the binding activity of NF-kappaB to the Fas promoter by RANKL, as detected by EMSA. In osteoclast precursors, the induction of Fas promoter activity by RANKL was dramatically reduced when NF-kappaB binding sites on the Fas promoter were mutated. CONCLUSION: RANKL upregulates Fas expression in osteoclast progenitors through NF-kappaB, making osteoclasts targets of Fas-stimulated apoptosis. In differentiated mature osteoclasts, RANKL reduces the levels of Fas expression and Fas-mediated apoptosis, acting as a survival factor. SN - 0884-0431 UR - https://www.unboundmedicine.com/medline/citation/15619676/RANKL_regulates_Fas_expression_and_Fas_mediated_apoptosis_in_osteoclasts_ DB - PRIME DP - Unbound Medicine ER -