Simvastatin inhibits osteoclast differentiation induced by bone morphogenetic protein-2 and RANKL through regulating MAPK, AKT and Src signaling

Misuzu Yamashita, Fumio Otsuka, Tomoyuki Mukai, Ryutaro Yamanaka, Hiroyuki Otani, Yoshinori Matsumoto, Eri Nakamura, Mariko Takano, Kenei Sada, Hirofumi Makino

Research output: Contribution to journalArticle

51 Citations (Scopus)

Abstract

The mevalonate pathway plays a crucial role in bone metabolism. Here we examined roles of simvastatin in osteoclast function and differentiation induced by RANKL and BMP-2 using mouse macrophage-like MLC-6 cells and human osteoclast precursor cells. MLC-6 cells expressed BMP type-I and -II receptors and Smads as well as osteoclast markers including TRAP, RANK, cathepsin-K, M-CSF receptor, MMP-9 and calcitonin receptor. Treatment with RANKL and BMP-2 acted synergistically to stimulate RANK, TRAP and cathepsin-K expression in MLC-6 cells. Simvastatin suppressed osteoclastic activity shown by increases in RANK, TRAP and cathepsin-K expression induced by RANKL and BMP-2. In contrast simvastatin alone had no effects on the osteoclastic markers in MLC-6 cells. Simvastatin activated ERK, SAPK/JNK and AKT pathways and inactivated Ras in MLC-6 cells. Simvastatin had no effect on BMP-induced Smad1/5/8 phosphorylation regardless of RANKL stimulation. Since chemical inhibition of ERK, SAPK/JNK and AKT increased TRAP and cathepsin-K expression induced by BMP-2 and RANKL, these pathways are functionally involved in inhibition of osteoclastic activity. In addition, Src phosphorylation induced by RANKL, which is involved in osteoclast differentiation, was suppressed by simvastatin. We further confirmed an inhibitory mechanism of simvastatin on osteoclast differentiation using human osteoclast precursor cells which express BMP receptor and Smad signaling machinery. Simvastatin also activated ERK pathways and inactivated Src phosphorylation in human osteoclasts differentiated by M-CSF and RANKL treatments. The inhibition of TRAP and RANK expression by simvastatin was reversed by ERK inhibition, whereas Src inhibitor enhanced simvastatin-induced suppression of osteoclast markers. Collectively, our data show that simvastatin inhibits osteoclastic differentiation through inhibiting Src as well as enhancing MAPK/AKT pathways.

Original languageEnglish
Pages (from-to)99-108
Number of pages10
JournalRegulatory Peptides
Volume162
Issue number1-3
DOIs
Publication statusPublished - Jun 2010

Fingerprint

Bone Morphogenetic Protein 2
Simvastatin
Osteoclasts
Cathepsin K
Phosphorylation
MAP Kinase Signaling System
Type I Bone Morphogenetic Protein Receptors
Type II Bone Morphogenetic Protein Receptors
Calcitonin Receptors
Macrophage Colony-Stimulating Factor Receptors
Bone Morphogenetic Protein Receptors
Mevalonic Acid
Macrophage Colony-Stimulating Factor
Macrophages
Matrix Metalloproteinases
Metabolism
Machinery
Bone

Keywords

  • Bone morphogenetic protein (BMP)
  • Mitogen-activated protein (MAP) kinase
  • MLC-6
  • Osteoclast
  • RANKL and statin

ASJC Scopus subject areas

  • Biochemistry
  • Clinical Biochemistry
  • Endocrinology
  • Physiology
  • Cellular and Molecular Neuroscience

Cite this

Simvastatin inhibits osteoclast differentiation induced by bone morphogenetic protein-2 and RANKL through regulating MAPK, AKT and Src signaling. / Yamashita, Misuzu; Otsuka, Fumio; Mukai, Tomoyuki; Yamanaka, Ryutaro; Otani, Hiroyuki; Matsumoto, Yoshinori; Nakamura, Eri; Takano, Mariko; Sada, Kenei; Makino, Hirofumi.

In: Regulatory Peptides, Vol. 162, No. 1-3, 06.2010, p. 99-108.

Research output: Contribution to journalArticle

@article{edbb71c89b9b4a078e1d9f97fec12baa,
title = "Simvastatin inhibits osteoclast differentiation induced by bone morphogenetic protein-2 and RANKL through regulating MAPK, AKT and Src signaling",
abstract = "The mevalonate pathway plays a crucial role in bone metabolism. Here we examined roles of simvastatin in osteoclast function and differentiation induced by RANKL and BMP-2 using mouse macrophage-like MLC-6 cells and human osteoclast precursor cells. MLC-6 cells expressed BMP type-I and -II receptors and Smads as well as osteoclast markers including TRAP, RANK, cathepsin-K, M-CSF receptor, MMP-9 and calcitonin receptor. Treatment with RANKL and BMP-2 acted synergistically to stimulate RANK, TRAP and cathepsin-K expression in MLC-6 cells. Simvastatin suppressed osteoclastic activity shown by increases in RANK, TRAP and cathepsin-K expression induced by RANKL and BMP-2. In contrast simvastatin alone had no effects on the osteoclastic markers in MLC-6 cells. Simvastatin activated ERK, SAPK/JNK and AKT pathways and inactivated Ras in MLC-6 cells. Simvastatin had no effect on BMP-induced Smad1/5/8 phosphorylation regardless of RANKL stimulation. Since chemical inhibition of ERK, SAPK/JNK and AKT increased TRAP and cathepsin-K expression induced by BMP-2 and RANKL, these pathways are functionally involved in inhibition of osteoclastic activity. In addition, Src phosphorylation induced by RANKL, which is involved in osteoclast differentiation, was suppressed by simvastatin. We further confirmed an inhibitory mechanism of simvastatin on osteoclast differentiation using human osteoclast precursor cells which express BMP receptor and Smad signaling machinery. Simvastatin also activated ERK pathways and inactivated Src phosphorylation in human osteoclasts differentiated by M-CSF and RANKL treatments. The inhibition of TRAP and RANK expression by simvastatin was reversed by ERK inhibition, whereas Src inhibitor enhanced simvastatin-induced suppression of osteoclast markers. Collectively, our data show that simvastatin inhibits osteoclastic differentiation through inhibiting Src as well as enhancing MAPK/AKT pathways.",
keywords = "Bone morphogenetic protein (BMP), Mitogen-activated protein (MAP) kinase, MLC-6, Osteoclast, RANKL and statin",
author = "Misuzu Yamashita and Fumio Otsuka and Tomoyuki Mukai and Ryutaro Yamanaka and Hiroyuki Otani and Yoshinori Matsumoto and Eri Nakamura and Mariko Takano and Kenei Sada and Hirofumi Makino",
year = "2010",
month = "6",
doi = "10.1016/j.regpep.2010.03.003",
language = "English",
volume = "162",
pages = "99--108",
journal = "Regulatory Peptides",
issn = "0167-0115",
publisher = "Elsevier",
number = "1-3",

}

TY - JOUR

T1 - Simvastatin inhibits osteoclast differentiation induced by bone morphogenetic protein-2 and RANKL through regulating MAPK, AKT and Src signaling

AU - Yamashita, Misuzu

AU - Otsuka, Fumio

AU - Mukai, Tomoyuki

AU - Yamanaka, Ryutaro

AU - Otani, Hiroyuki

AU - Matsumoto, Yoshinori

AU - Nakamura, Eri

AU - Takano, Mariko

AU - Sada, Kenei

AU - Makino, Hirofumi

PY - 2010/6

Y1 - 2010/6

N2 - The mevalonate pathway plays a crucial role in bone metabolism. Here we examined roles of simvastatin in osteoclast function and differentiation induced by RANKL and BMP-2 using mouse macrophage-like MLC-6 cells and human osteoclast precursor cells. MLC-6 cells expressed BMP type-I and -II receptors and Smads as well as osteoclast markers including TRAP, RANK, cathepsin-K, M-CSF receptor, MMP-9 and calcitonin receptor. Treatment with RANKL and BMP-2 acted synergistically to stimulate RANK, TRAP and cathepsin-K expression in MLC-6 cells. Simvastatin suppressed osteoclastic activity shown by increases in RANK, TRAP and cathepsin-K expression induced by RANKL and BMP-2. In contrast simvastatin alone had no effects on the osteoclastic markers in MLC-6 cells. Simvastatin activated ERK, SAPK/JNK and AKT pathways and inactivated Ras in MLC-6 cells. Simvastatin had no effect on BMP-induced Smad1/5/8 phosphorylation regardless of RANKL stimulation. Since chemical inhibition of ERK, SAPK/JNK and AKT increased TRAP and cathepsin-K expression induced by BMP-2 and RANKL, these pathways are functionally involved in inhibition of osteoclastic activity. In addition, Src phosphorylation induced by RANKL, which is involved in osteoclast differentiation, was suppressed by simvastatin. We further confirmed an inhibitory mechanism of simvastatin on osteoclast differentiation using human osteoclast precursor cells which express BMP receptor and Smad signaling machinery. Simvastatin also activated ERK pathways and inactivated Src phosphorylation in human osteoclasts differentiated by M-CSF and RANKL treatments. The inhibition of TRAP and RANK expression by simvastatin was reversed by ERK inhibition, whereas Src inhibitor enhanced simvastatin-induced suppression of osteoclast markers. Collectively, our data show that simvastatin inhibits osteoclastic differentiation through inhibiting Src as well as enhancing MAPK/AKT pathways.

AB - The mevalonate pathway plays a crucial role in bone metabolism. Here we examined roles of simvastatin in osteoclast function and differentiation induced by RANKL and BMP-2 using mouse macrophage-like MLC-6 cells and human osteoclast precursor cells. MLC-6 cells expressed BMP type-I and -II receptors and Smads as well as osteoclast markers including TRAP, RANK, cathepsin-K, M-CSF receptor, MMP-9 and calcitonin receptor. Treatment with RANKL and BMP-2 acted synergistically to stimulate RANK, TRAP and cathepsin-K expression in MLC-6 cells. Simvastatin suppressed osteoclastic activity shown by increases in RANK, TRAP and cathepsin-K expression induced by RANKL and BMP-2. In contrast simvastatin alone had no effects on the osteoclastic markers in MLC-6 cells. Simvastatin activated ERK, SAPK/JNK and AKT pathways and inactivated Ras in MLC-6 cells. Simvastatin had no effect on BMP-induced Smad1/5/8 phosphorylation regardless of RANKL stimulation. Since chemical inhibition of ERK, SAPK/JNK and AKT increased TRAP and cathepsin-K expression induced by BMP-2 and RANKL, these pathways are functionally involved in inhibition of osteoclastic activity. In addition, Src phosphorylation induced by RANKL, which is involved in osteoclast differentiation, was suppressed by simvastatin. We further confirmed an inhibitory mechanism of simvastatin on osteoclast differentiation using human osteoclast precursor cells which express BMP receptor and Smad signaling machinery. Simvastatin also activated ERK pathways and inactivated Src phosphorylation in human osteoclasts differentiated by M-CSF and RANKL treatments. The inhibition of TRAP and RANK expression by simvastatin was reversed by ERK inhibition, whereas Src inhibitor enhanced simvastatin-induced suppression of osteoclast markers. Collectively, our data show that simvastatin inhibits osteoclastic differentiation through inhibiting Src as well as enhancing MAPK/AKT pathways.

KW - Bone morphogenetic protein (BMP)

KW - Mitogen-activated protein (MAP) kinase

KW - MLC-6

KW - Osteoclast

KW - RANKL and statin

UR - http://www.scopus.com/inward/record.url?scp=77952671313&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=77952671313&partnerID=8YFLogxK

U2 - 10.1016/j.regpep.2010.03.003

DO - 10.1016/j.regpep.2010.03.003

M3 - Article

VL - 162

SP - 99

EP - 108

JO - Regulatory Peptides

JF - Regulatory Peptides

SN - 0167-0115

IS - 1-3

ER -