Ok-Joo Sul

Saturated fatty acids enhance osteoclast survival.

Hyperlipidemia and marrow fat are associated with lowering bone density in vivo, suggesting that lipid contributes to bone loss. Using bone marrow-derived macrophages, we investigated the effect of saturated fatty acids (SFA) on osteoclastogenesis. The level of free fatty acids and adiposity in bone marrow was significantly elevated in obese mice. SFA increased osteoclast (OC) survival by preventing apoptosis. SFA caused the production of MIP-1alpha and led to activation of nuclear factor (NF)-kappaB in the OC. The absence of Toll-like receptor 4 (TLR4) or myeloid differentiation factor 88 (MyD88) abolished the survival effect of SFA on OC.

Gold Nanoparticles Inhibited the Receptor Activator of Nuclear Factor-κB Ligand (RANKL)-Induced Osteoclast Formation by Acting as an Antioxidant

Gold nanoparticles inhibited osteoclast (OC) formation induced by the receptor activator of nuclear factor-κB ligand (RANKL) in bone marrow-derived macrophages (BMMs). This was accompanied by a decreased level of tartrate-resistant alkaline phosphatase (TRAP) and less activation of nuclear factor (NF)-κB. The nanoparticles also reduced the production of reactive oxygen species (ROS) in response to RANKL and upregulated RANKL-induced glutathione peroxidase-1 (Gpx-1), suggesting a role as an antioxidant in the BMM. The inhibitory effects on OC formation might have been due to elevated defense against oxidative stress.

Curcumin protects against ovariectomy‐induced bone loss and decreases osteoclastogenesis

Curcumin has anti‐oxidative activity. In view of the increasing evidence for a biochemical link between increased oxidative stress and reduced bone density we hypothesized that curcumin might increase bone density by elevating antioxidant activity in some target cell type. We measured bone density by Micro‐CT, enzyme expression levels by quantitative PCR or enzyme activity, and osteoclast (OC) formation by tartrate‐resistant acid phosphatase staining. The bone mineral density of the femurs of curcumin‐administered mice was significantly higher than that of vehicle‐treated mice after ovariectomy (OVX) and this was accompanied by reduced amounts of serum collagen‐type I fragments, which are markers of bone resorption. Curcumin suppressed OC formation by increasing receptor activator of nuclear factor‐κB ligand (RANKL)‐induced glutathione peroxidase‐1, and reversed the stimulatory effect of homocysteine, a known H2O2 generator, on OC formation by restoring Gpx activity. Curcumin generated an aberrant RANKL signal characterized by reduced expression of nuclear factor of activated T cells 2 (NFAT2) and attenuated activation of mitogen‐activated protein kinases (ERK, JNK, and p38). Curcumin thus inhibited OVX‐induced bone loss, at least in part by reducing osteoclastogenesis as a result of increased antioxidant activity and impaired RANKL signaling. These findings suggest that bone loss associated with estrogen deficiency could be attenuated by curcumin administration. J. Cell. Biochem. 112: 3159–3166, 2011.

Absence of MCP-1 leads to elevated bone mass via impaired actin ring formation.

Monocyte chemoattractant protein‐1 (MCP‐1) is associated with various inflammatory diseases involving bone loss, and is expressed along with its receptor by bone marrow‐derived macrophages (BMM), which are osteoclast (OC) precursors. To investigate the role of MCP‐1 in bone remodeling, we compared MCP‐1‐knockout (KO) mice with wild‐type (WT) mice. The absence of MCP‐1 increased bone mass and lowered serum collagen type I fragments (CTX‐1) and TRACP 5b, but had no significant effect on the N‐terminal propeptide of type I procollagen, suggesting that OCs are primarily responsible for the bone phenotype observed in the absence of MCP‐1. MCP‐1 deficiency resulted in reduced numbers and activity of OCs in vitro. It also led to a reduced level of c‐Fms and receptor activator of nuclear factor‐κB receptor and impaired actin ring formation. Activation of ERK, Akt, Rac1, and Rho upon M‐CSF stimulation was also reduced and our evidence suggests that the aberrant actin ring formation was partly due to reduced activation of these molecules. Our findings point to a role of osteoclast MCP‐1 in regulating bone remodeling. The higher bone mass in the femurs of MCP‐1‐KO mice could be, at least in part, due to decreased osteoclastogenesis and bone resorption resulting from aberrant M‐CSF signaling in OCs. J. Cell. Physiol. 227: 1619–1627, 2012.

Monocyte Chemoattractant Protein-1 Deficiency Attenuates Oxidative Stress and Protects against Ovariectomy-Induced Chronic Inflammation in Mice

Background Loss of ovarian function is highly associated with an elevated risk of metabolic disease. Monocyte chemoattractant protein-1 (MCP-1, C-C chemokine ligand 2) plays critical roles in the development of inflammation, but its role in ovariectomy (OVX)-induced metabolic disturbance has not been known. Methodology and Principal Findings We investigated the role of MCP-1 in OVX-induced metabolic perturbation using MCP-1-knockout mice. OVX increased fat mass, serum levels of MCP-1, macrophage-colony stimulating factor (M-CSF), and reactive oxygen species (ROS), whereas MCP-1 deficiency attenuated these. OVX-induced increases of visceral fat resulted in elevated levels of highly inflammatory CD11c-expressing cells as well as other immune cells in adipose tissue, whereas a lack of MCP-1 significantly reduced all of these levels. MCP-1 deficiency attenuated activation of phospholipase Cγ2, transforming oncogene from Ak strain, and extracellular signal-regulated kinase as well as generation of ROS, which is required for up-regulating CD11c expression upon M-CSF stimulation in bone marrow-derived macrophages. Conclusions/Significance Our data suggested that MCP-1 plays a key role in developing metabolic perturbation caused by a loss of ovarian functions through elevating CD11c expression via ROS generation.

Increased fat due to estrogen deficiency induces bone loss by elevating monocyte chemoattractant protein-1 (MCP-1) production.

Ovariectomy (OVX)-induced estrogen withdrawal resulted in both bone loss and an increase in fat. We observed elevated osteoclast (OC) formation by bone marrow-derived macrophages treated with medium conditioned by fats from OVX mice, but not from sham-operated mice. Fats from OVX mice expressed and secreted higher levels of monocyte chemoattractant protein-1 (MCP-1) than those from sham-operated mice. Increased fat resulting from estrogen deficiency is thus responsible for bone loss due to enhanced OC formation, which is, at least partly, a consequence of elevated MCP-1 production.

Hemeoxygenase-1 maintains bone mass via attenuating a redox imbalance in osteoclast.

Heme oxygenase-1 (HO-1) has long been considered to be an endogenous antioxidant. However, the role of HO-1 is highly controversial in developing metabolic diseases. We hypothesized that HO-1 plays a role in maintaining bone mass by alleviating a redox imbalance. We investigated its role in bone remodeling. The absence of HO-1 in mice led to decreased bone mass with elevated activity and number of OCs, as well as higher serum levels of reactive oxygen species (ROS). HO-1, which is constitutively expressed at a high level in osteoclast (OC) precursors, was down-regulated during OC differentiation. HO-1 deficiency in bone marrow macrophages (BMM) in vitro resulted in increased numbers and activity of OCs due to enhanced receptor activator of nuclear factor-κB ligand (RANKL) signaling. This was associated with increased activation of nuclear factor-κB and of nuclear factor of activated T-cells, cytoplasmic 1 along with elevated levels of intracellular calcium and ROS. Decreased bone mass in the absence of HO-1 appears to be mainly due to increased osteoclastogenesis and bone resorption resulting from elevated RANKL signaling in OCs. Our data highlight the potential role of HO-1 in maintaining bone mass by negatively regulating OCs.

TNFRSF14 deficiency protects against ovariectomy-induced adipose tissue inflammation

To elucidate the role of tumor necrosis factor receptor superfamily member 14 (TNFRSF14) in metabolic disturbance due to loss of ovarian function, ovariectomy (OVX) was performed in TNFRSF 14-knockout mice. OVX increased fat mass and infiltration of highly inflammatory CD11c cells in the adipose tissue (AT), which was analyzed by flow cytometry, and resulted in disturbance of glucose metabolism, whereas TNFRSF14 deficiency attenuated these effects. TNFRSF14 deficiency decreased recruitment of CD11c-expressing cells in AT and reduced the polarization of bone marrow-derived macrophages to M1. Upon engagement of LIGHT, a TNFRSF14 ligand, TNFRSF14 enhanced the expression of CD11c via generation of reactive oxygen species, suggesting a role of TNFRSF14 as a redox modulator. TNFRSF14 participated in OVX-induced AT inflammation via upregulation of CD11c, resulting in metabolic perturbation. TNFRSF14 could be used as a therapeutic target for the treatment of postmenopausal syndrome by reducing AT inflammation.

Reactive oxygen species induce the association of SHP-1 with c-Src and the oxidation of both to enhance osteoclast survival

Loss of ovarian function causes oxidative stress as well as bone loss. We hypothesized that reactive oxygen species (ROS) induced by the failure of ovarian function are responsible for the bone loss by increasing the number of osteoclasts (OC). We found that ROS enhanced OC survival via Src homology 2 domain-containing phosphatase-1 (SHP-1), c-Src, Akt, and ERK. ROS induced the association of SHP-1 with c-Src as well as the oxidation of c-Src and SHP-1. This resulted in inactivation of SHP-1 and activation of c-Src via phosphorylation of Tyr(416). Knockdown of c-Src or SHP-1 abolished the effect of ROS on OC survival. Moreover, downregulation of SHP-1 upregulated activation of c-Src, Akt, and ERK in the absence of any stimulus, suggesting that inactivation of SHP-1 is required for OC survival. We demonstrated that the association and oxidation of c-Src and SHP-1 by ROS are key steps in enhancing OC survival, which are responsible for increased bone loss when ovarian function ceases.

Carbon monoxide protects against ovariectomy-induced bone loss by inhibiting osteoclastogenesis

Carbon monoxide (CO) has been shown to have remarkable therapeutic value at low dosage by suppressing inflammation via inhibitory effects on macrophages, which are also precursors of osteoclasts (OC). The objective of the present study was to determine whether CO limits bone loss through its effects on osteoclastogenesis. Intraperitoneal injection of CO-releasing molecule 2 (CORM2) into mice with reduced bone mass due to ovariectomy (OVX) resulted in significantly elevated bone mass. Increased serum levels of collagen-type I fragments, tartrate-resistant acid phosphatase 5b, and reactive oxygen species (ROS) due to OVX were also decreased when treated with CORM2. In vitro, CORM2 inhibited receptor activator of nuclear factor-κB ligand (RANKL)-induced OC formation without affecting bone resorption. CORM2 reduced long-lasting ROS levels and nuclear factor-κB (NF-κB) activation in response to RANKL. Inhibition of NADPH oxidase partially reduced the inhibitory effect of CO. CO induced increase of peroxiredoxin 1 (PRX1) in BMM. Down-regulation of PRX1 reduced the inhibitory effect of CO on OC formation and sustained the ROS levels induced by RANKL, suggesting that CO reduces generation of ROS and scavenges ROS to inhibit osteoclastogenesis. These data suggest that the inhibitory effect of CO on osteoclastogenesis is caused by impaired RANKL signaling through defective NF-κB activation and reduced levels of long-lasting ROS. These changes result in decreased bone loss. Our data highlight the potential utility of CO for ameliorating bone loss induced by loss of ovarian function.