Activation of Liver X Receptor (LXR) Inhibits Receptor Activator of Nuclear Factor κB Ligand (RANKL)-induced Osteoclast Differentiation in an LXRβ-dependent Mechanism

Abstract

Bone destruction is the major pathological process in many bone metabolic diseases and is a result of increased osteoclast formation and bone resorption. The liver X receptors (␣,␤), important regulators of cholesterol metabolism and inflammatory signaling, have recently been observed to play a role in both physiological and pathological bone turnover. However, the relationship between liver X receptors (LXR) and osteoclast differentiation/formation remains unknown. Here, we report that the LXR ligand GW3965 is able to clearly and potently inhibit the formation of mature osteoclasts from receptor activator of nuclear factor B ligand (RANKL)-stimulated human and murine osteoclast precursors. This results in a significant inhibition of bone resorption. We observed that GW3965 significantly inhibited expression of the osteoclast markers tartrateresistant acid phosphatase, cathepsin K, osteoclast-associated receptor (OSCAR), and calcitonin receptor, appearing to act in an NFATc1/p38/microphthalmia-associated transcription factor (MITF)-dependent mechanism, independently of receptor activator of nuclear factor B or c-Fos and not directly involving the NFB pathways. GW3965 was less effective in RAW264.7 monocyte/macrophage cells, which are more committed into the osteoclast lineage. Also, GW3965 seemed to act differently depending on the source of the progenitor cells as it had no effect on calvarial osteoclasts, compared with marrow or bloodderived monocytes. As these effects were abolished in osteoclast precursors derived from LXR␤ ؊/؊ mice, we suggest that GW3965 acts via an LXR␤-dependent mechanism. Taken together, our results suggest that the LXR can act as an important inhibitor of RANKL-mediated osteoclast differentiation.Bone remodeling occurs continuously throughout life to maintain bone quality in response to mechanical stress and hormonal regulation. To retain a normal bone mass, bone is resorbed by osteoclasts, and new bone matrix is deposited by osteoblasts in a tightly coupled process. Imbalances in this process can lead to excessive resorption by the osteoclasts and increased bone fragility, as observed in many common skeletal diseases (osteoporosis, metastatic bone disease, and Paget disease of bone) (1, 2). Osteoclasts are multinucleated giant cells derived from the pluripotent hematopoietic stem cell lineage. Differentiation of the precursor cells into the osteoclast lineage is principally regulated by macrophage colony-stimulating factor (M-CSF) and receptor activator of NFB ligand (RANKL) 2 (3). Binding of RANKL to its receptor RANK activates different signaling cascades (NFB, MAPK, and Ca 2ϩ oscillation/calcineurin) to induce nuclear factor of activated T cells c1 (NFATc1), the key transcription factor in osteoclastogenesis (4, 5). NFATc1 in turn induces the transcription of osteoclast-specific genes, including tartrate-resistant acid phosphatase (TRAP/acp5), cathepsin K (ctsk), osteoclast-associated receptor (oscar), and calcitonin receptor (calcr) (6).The liver X receptors, LXR␣ and LXR␤, are me...

Liver X receptors orchestrate osteoblast/osteoclast crosstalk and counteract pathologic bone loss

“…Experiments with LXR aÀ/À, LXR bÀ/À, and LXR a/b-doubledeficient cells revealed that this LXR ligand-induced block of osteoclast differentiation involved LXR-mediated effects in osteoblasts and osteoclasts, although expression of LXRs in osteoblasts was essential for the observed effects. These inhibitory effects of LXRs on osteoclastogenesis is in accordance with a recent study by Remen and colleagues, (13) which reported direct inhibitory effects of LXR ligands on osteoclasts. In this study the authors observed a direct inhibition of the end-stage differentiation of osteoclasts by LXR ligands.…”

“…In the absence of osteoblasts, we observed no effect of the LXR ligand on osteoclast numbers, although the osteoclasts were reduced in size and activity. This slight discrepancy might be explained by the fact that the study by Remen and colleagues (13) focused on later phases of osteoclastogenesis and used rather high concentrations of LXR ligands. In contrast, we added the LXR ligand at an early time point and focused on concentrations at a range between 10 nM and 1 mM.…”

“…These alternative mechanisms might include the modulation of other osteoblast-derived factors such as semaphorins, which influence osteoclastogenesis, (20) as well as the observed direct inhibitory effects of LXR agonists on osteoclast differentiation. (13) Although we observed an LXR-mediated inhibition of RANKL expression in vitro and in vivo, future studies are necessary to elucidate the exact underlying molecular mechanisms. Like other nuclear receptors, LXRs are able to recruit co-repressors in a promoter-specific manner and thereby silences transcriptional activation of distinct target genes.…”

“…(12) Nevertheless, recent in vitro data indicate a direct inhibitory effect of LXR ligands on the end-stage differentiation of osteoclasts and on the expression of bone formation markers in primary osteoblasts. (12,13) Together these data raise the question of the precise role of LXRs in osteoblast and osteoclast function and during bone turnover in vivo.…”

An ELIXIR for bone loss?

“…The same authors went on to show that an LXR agonist suppressed in vitro osteoclast formation from bone marrow macrophages (BMMs) exposed to the key osteoclastogenic cytokines macrophage colony-stimulating factor (M-CSF) and RANKL via a mechanism involving blockade of RANKL-induced signaling. (27) The current authors also had access to mice carrying all three mutant LXR genotypes, but chose to use the animals mainly as a source of osteoblasts and BMMs for in vitro studies. The two cell types were co-cultured in the presence of an LXR agonist or parathyroid hormone, which stimulates osteoclastogenesis in this setting by enhancing production of M-CSF and RANKL by the mesenchymal cells.…”