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Punicalagin attenuates osteoclast differentiation by impairing NFATc1 expression and blocking Akt- and JNK-dependent pathways

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Abstract

Punicalagin is a bioactive polyphenol that is classified as an ellagitannin. Although punicalagin has been shown to have various pharmacological effects, such as anti-oxidative, anti-inflammatory, and anti-tumor effects, no studies have reported the effects of punicalagin on osteoclasts (OCLs). In this study, we investigated the effects of punicalagin on OCL differentiation by receptor activator of nuclear factor kappa-B ligand in the murine monocytic RAW-D cell line and bone marrow-derived macrophages (BMMs). Treatment with punicalagin significantly inhibited OCL formation from RAW-D cells and BMMs and prevented bone resorption of BMM-derived OCLs. Moreover, punicalagin impaired multinucleation and actin-ring formation in OCLs, and decreased the protein levels of nuclear factor of activated T cells cytoplasmic-1 (NFATc1), which is a master regulator of OCL differentiation, and concomitantly reduced the expression levels of Src and cathepsin K, which are transcriptionally regulated by NFATc1. The effects of punicalagin on intracellular signaling during the OCL differentiation of BMMs indicated that punicalagin-treated OCLs displayed markedly reduced phosphorylation of Jun N-terminal kinase and Akt, and partially impaired phosphorylation of extracellular signal-regulated kinase, p38 mitogen-activated protein kinase, and inhibitor of nuclear factor kappa-B alpha compared with untreated OCLs. Thus, punicalagin may affect bone metabolism by inhibiting OCL differentiation.

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Abbreviations

OCLs:

Osteoclasts

RANKL:

Receptor activator of nuclear factor kappa-B ligand

BMMs:

Bone marrow-derived macrophages

NFATc1:

Nuclear factor of activated T cells cytoplasmic-1

IκBα:

Nuclear factor kappa-B alpha

TRAP:

Tartrate-resistant acid phosphatase

M-CSF:

Macrophage colony-stimulating factor

NF-κB:

Nuclear factor kappa-B

PI3 K:

Phosphatidylinositol 3-kinase

JNK:

Jun N-terminal kinase

Erk:

Extracellular signal-regulated kinase

MAPK:

Mitogen-activated protein kinase

HO-1:

Heme oxygenase-1

NFAT:

Nuclear factor of activated T cells

Abs:

Antibodies

SD:

Standard deviations

PMSF:

Phenylmethylsulfonyl fluoride

TBS-T:

Tris buffered saline with 0.1 % Tween 20

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Acknowledgments

We thank Dr. Kazuhisa Nishishita for providing recombinant RANKL, and Dr. Eiko Sakai for useful comments. This study was supported in part by Grants-in-Aid for Scientific Research (B) Grant Numbers 25293383, 15H05298, and for Exploratory Research Grant Number 30264055 (T.T).

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Authors and Affiliations

  1. Division of Dental Pharmacology, Nagasaki University Graduate School of Biomedical Sciences, Sakamoto 1-7-1, Nagasaki, 852-8588, Japan

    Mayumi Iwatake, Kuniaki Okamoto & Takayuki Tsukuba

  2. Division of Natural Product Chemistry, Nagasaki University Graduate School of Biomedical Sciences, Bunkyo, 1-14, Nagasaki, 852-8521, Japan

    Takashi Tanaka

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  1. Mayumi Iwatake
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  2. Kuniaki Okamoto
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  3. Takashi Tanaka
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  4. Takayuki Tsukuba
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Correspondence to Takayuki Tsukuba.

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Iwatake, M., Okamoto, K., Tanaka, T. et al. Punicalagin attenuates osteoclast differentiation by impairing NFATc1 expression and blocking Akt- and JNK-dependent pathways. Mol Cell Biochem 407, 161–172 (2015). https://doi.org/10.1007/s11010-015-2466-3

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