Impact of Intolerance of Uncertainty on Brain Structural Changes in Panic Disorder.

OBJECTIVE: This study investigated the impact of intolerance of uncertainty (IU) on structural changes in the brain and symptom severity in patients with panic disorder. METHODS: This study included 90 participants diagnosed with panic disorder. The IU Scale, Panic Disorder Severity Scale (PDSS), Beck Depression Inventory-II (BDI-II), Penn State Worry Questionnaire (PSWQ), Self-Forgiveness Scale (SFS), and Short Form 36 Health Survey (SF) were used. A voxel-wise correlation analysis was conducted to investigate the structural differences in the gray matter. RESULTS: As IU increased, the cortical thickness of the right lingual gyrus decreased significantly, while the gray matter volume of the right pars triangularis increased. The cortical thickness of the right lingual gyrus showed a significant negative correlation with the BDI-II score and a positive correlation with the SFS. Additionally, the gray matter volume of the right pars triangularis was positively correlated with the PDSS, PSWQ, and BDI-II scores and negatively correlated with the mental health domain of the SF. CONCLUSION: According to our findings, elevated IU in participants with panic disorder was associated with cortical thinning in the lingual gyrus and increased gray matter volume in the pars triangularis. These structural alterations may also have an impact on perceived quality of life, as well as high levels of depression and anxiety.

Harpagoside attenuates local bone Erosion and systemic osteoporosis in collagen-induced arthritis in mice.

BACKGROUND: Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease that causes local bone erosion and systemic osteoporosis. Harpagoside (HAR), an iridoid glycoside, has various pharmacological effects on pain, arthritis, and inflammation. Our previous study suggests that HAR is more deeply involved in the mechanism of bone loss caused by inflammatory stimuli than hormonal changes. Here, we identified the local and systemic bone loss inhibitory effects of HAR on RA and its intracellular mechanisms using a type 2 collagen-induced arthritis (CIA) mouse model. METHODS: The anti-osteoporosis and anti-arthritic effects of HAR were evaluated on bone marrow macrophage in vitro and CIA in mice in vivo by obtaining clinical scores, measuring hind paw thickness and inflammatory cytokine levels, micro-CT and histopathological assessments, and cell-based assay. RESULTS: HAR markedly reduced the clinical score and incidence rate of CIA in both the prevention and therapy groups. Histological analysis demonstrated that HAR locally ameliorated the destruction of bone and cartilage and the formation of pannus. In this process, HAR decreased the expression of inflammatory cytokines, such as tumor necrosis factor-α, interleukin (IL)-6, and IL-1ß in the serum of CIA mice. Additionally, HAR downregulated the expression of receptor activator of nuclear factor-κB ligand and upregulated that of osteoprotegerin. HAR suppressed systemic bone loss by inhibiting osteoclast differentiation and osteoclast marker gene expression in a CIA mouse model. CONCLUSIONS: Taken together, these findings show the beneficial effect of HAR on local symptoms and systemic bone erosion triggered by inflammatory arthritis.

Quantitative Photothermal Characterization with Bioprinted 3D Complex Tissue Constructs for Early-Stage Breast Cancer Therapy Using Gold Nanorods.

Plasmonic photothermal therapy (PPTT) using gold nanoparticles (AuNPs) has shown great potential for use in selective tumor treatment, because the AuNPs can generate destructive heat preferentially upon irradiation. However, PPTT using AuNPs has not been added to practice, owing to insufficient heating methods and tissue temperature measurement techniques, leading to unreliable and inaccurate treatments. Because the photothermal properties of AuNPs vary with laser power, particle optical density, and tissue depth, the accurate prediction of heat generation is indispensable for clinical treatment. In this report, bioprinted 3D complex tissue constructs comprising processed gel obtained from porcine skin and human decellularized adipose tissue are presented for characterization of the photothermal properties of gold nanorods (AuNRs) having an aspect ratio of 3.7 irradiated by a near-infrared laser. Moreover, an analytical function is suggested for achieving PPTT that can cause thermal damage selectively on early-stage human breast cancer by regulating the heat generation of the AuNRs in the tissue.

Nicotinamide phosphoribosyltransferase inhibits receptor activator of nuclear factor-κB ligand-induced osteoclast differentiation in vitro.

The adipokine nicotinamide phosphoribosyltransferase (Nampt), also known as pre-B-cell colony-enhancing factor or the insulin-mimetic hormone visfatin, has a crucial role in the conversion of nicotinamide to nicotinamide mononucleotide during biosynthesis of the coenzyme nicotinamide adenine dinucleotide. Previous reports have demonstrated the inhibitory effects of Nampt on osteoclast formation from human peripheral blood mononuclear cells and CD14+ monocytes. However, the effects of Nampt on bone marrow macrophage (BMM)­derived osteoclastogenesis and its precise role in the process remain unclear. The present in vitro study used recombinant Nampt and BMMs as osteoclast precursors demonstrated that Nampt suppresses receptor activator of nuclear factor­κB ligand (RANKL)­induced osteoclastogenesis by decreasing the phosphorylation of various early signal transducers, including c­Jun N­terminal kinase, Akt, glycogen synthase kinase­3 ß, Bruton's tyrosine kinase and phospholipase C γ­2. In addition, western blotting and reverse transcription­quantitative polymerase chain reaction analysis indicated that Nampt downregulates the mRNA and protein expression levels of c­Fos and nuclear factor of activated T cells, cytoplasmic 1, leading to a decrease in the expression of osteoclast­specific genes including tartrate­resistant acid phosphatase, osteoclast­associated receptor and cathepsin K. However, the bone­resorbing activity of mature osteoclasts treated with Nampt was similar to untreated control osteoclasts. This finding indicates that Nampt exerts its anti­osteoclastogenic activity by targeting osteoclast precursor cells rather than mature osteoclasts. Consequently, the present study demonstrated that Nampt acts as a negative regulator of RANKL­mediated differentiation of BMMs into osteoclasts, suggesting the potential therapeutic targets to treat bone-related disorders such as osteoporosis.

Ethanolic extract of Schizonepeta tenuifolia attenuates osteoclast formation and activation in vitro and protects against lipopolysaccharide-induced bone loss in vivo.

BACKGROUND: Excessive osteoclast activity is a major cause of metabolic bone disorders, such as osteopenia, rheumatoid arthritis, and osteoporosis. Thus, discovery of agents targeting osteoclast differentiation and bone resorption is important for development of novel treatments for bone diseases. It has been demonstrated that ethanolic extract of schizonepeta tenuifolia (EEST) has potent anti-oxidant and anti-inflammatory activities. However, the beneficial effects of EEST on bone metabolism have not been studied. Therefore, we intend to investigate the effects of EEST on osteoclast differentiation. METHODS: We examined the effects and mechanisms of action of the EEST on osteoclastogenesis in vitro in bone marrow macrophages (BMMs) stimulated with receptor activator of nuclear factor kappa-B ligand (RANKL) and in vivo using a mouse model of lipopolysaccharide (LPS)-induced bone destruction. RESULTS: We found that EEST inhibited phosphorylation of Akt and IkB at early stages of RANKL-induced osteoclastogenesis. Furthermore, EEST negatively controlled the transcription and translation levels of nuclear factor of activated T cells c1 (NFATc1) and the translation level of c-Fos at the final stage of osteoclast differentiation. Reflecting these effects, EEST blocked both filamentous actin (F-actin) ring formation and bone resorbing activity of mature osteoclasts in vitro. The inhibitory effects of EEST on osteoclast formation and activity were observed in an LPS-mediated bone erosion mouse model using micro-CT and histological analysis. CONCLUSIONS: EEST is a potential agent that is able to treat osteoclast-related bone diseases, such as osteoporosis.

Dendrobium moniliforme Exerts Inhibitory Effects on Both Receptor Activator of Nuclear Factor Kappa-B Ligand-Mediated Osteoclast Differentiation in Vitro and Lipopolysaccharide-Induced Bone Erosion in Vivo.

Dendrobium moniliforme (DM) is a well-known plant-derived extract that is widely used in Oriental medicine. DM and its chemical constituents have been reported to have a variety of pharmacological effects, including anti-oxidative, anti-inflammatory, and anti-tumor activities; however, no reports discuss the beneficial effects of DM on bone diseases such as osteoporosis. Thus, we investigated the relationship between DM and osteoclasts, cells that function in bone resorption. We found that DM significantly reduced receptor activator of nuclear factor kappa-B ligand (RANKL)-induced tartrate-resistant acid phosphatase (TRAP)-positive osteoclast formation; DM directly induced the down-regulation of c-Fos and nuclear factor of activated T cells c1 (NFATc1) without affecting other RANKL-dependent transduction pathways. In the later stages of osteoclast maturation, DM negatively regulated the organization of filamentous actin (F-actin), resulting in impaired bone-resorbing activity by the mature osteoclasts. In addition, micro-computed tomography (µ-CT) analysis of the murine model revealed that DM had a beneficial effect on lipopolysaccharide (LPS)-mediated bone erosion. Histological analysis showed that DM attenuated the degradation of trabecular bone matrix and formation of TRAP-positive osteoclasts in bone tissues. These results suggest that DM is a potential candidate for the treatment of metabolic bone disorders such as osteoporosis.

Protocatechuic Acid Attenuates Osteoclastogenesis by Downregulating JNK/c-Fos/NFATc1 Signaling and Prevents Inflammatory Bone Loss in Mice.

Protocatechuic acid (PCA) plays a critical role in nutritional metabolism; it is a major metabolite of anthocyanins, which are flavonoids with a range of health benefits. PCA has a variety of biological activities including anti-oxidant, antiinflammatory, anti-apoptosis, and anti-microbial activities. However, the pharmacological effect of PCA, especially on osteoclastogenesis, remains unknown. We examined the effect of PCA on receptor activator of NF-κB ligand (RANKL)-induced osteoclast differentiation and bone resorption. PCA dose-dependently inhibited RANKL-induced osteoclast differentiation in mouse bone marrow macrophages (BMMs) and suppressed the bone-resorbing activity of mature osteoclasts. At the molecular level, PCA suppressed RANKL-induced phosphorylation of JNK among MAPKs only, without significantly affecting the early signaling pathway. PCA also suppressed RANKL-stimulated expression of c-Fos and nuclear factor of activated T cells c1 (NFATc1) at the mRNA and protein levels, without altering c-Fos mRNA expression. Additionally, PCA down-regulated the expression of downstream osteoclastogenesis-related genes including ß3-integrin, DC-STAMP, OC-STAMP, Atp6v0d2, CTR, and CtsK. Mice treated with PCA efficiently recovered from lipopolysaccharide-induced bone loss in vivo. Thus, PCA inhibits RANKL-induced osteoclast differentiation and function by suppressing JNK signaling, c-Fos stability, and expression of osteoclastic marker genes. These results suggest that PCA could be useful in treatment of inflammatory bone disorders.

Niclosamide suppresses RANKL-induced osteoclastogenesis and prevents LPS-induced bone loss.

Niclosamide (5-chloro-salicyl-(2-chloro-4-nitro) anilide) is an oral anthelmintic drug used for treating intestinal infection of most tapeworms. Recently, niclosamide was shown to have considerable efficacy against some tumor cell lines, including colorectal, prostate, and breast cancers, and acute myelogenous leukemia. Specifically, the drug was identified as a potent inhibitor of signal transducer and activator of transcription 3 (STAT3), which is associated with osteoclast differentiation and function. In this study, we assessed the effect of niclosamide on osteoclastogenesis in vitro and in vivo. Our in vitro study showed that receptor activator of nuclear factor-kappaB ligand (RANKL)-induced osteoclast differentiation was inhibited by niclosamide, due to inhibition of serine-threonine protein kinase (Akt) phosphorylation, inhibitor of nuclear factor-kappaB (IκB), and STAT3 serine(727). Niclosamide decreased the expression of the major transcription factors c-Fos and NFATc1, and thereafter abrogated the mRNA expression of osteoclast-specific genes, including TRAP, OSCAR, αv/ß3 integrin (integrin αv, integrin ß3), and cathepsin K (CtsK). In an in vivo model, niclosamide prevented lipopolysaccharide-induced bone loss by diminishing osteoclast activity. Taken together, our results show that niclosamide is effective in suppressing osteoclastogenesis and may be considered as a new and safe therapeutic candidate for the clinical treatment of osteoclast-related diseases such as osteoporosis.

WHI-131 Promotes Osteoblast Differentiation and Prevents Osteoclast Formation and Resorption in Mice.

The small molecule WHI-131 is a potent therapeutic agent with anti-inflammatory, antiallergic, and antileukemic potential. However, the regulatory effects of WHI-131 on osteoblast and osteoclast activity are unclear. We examined the effects of WHI-131 on osteoblast and osteoclast differentiation with respect to bone remodeling. The production of receptor activator of nuclear factor kappa-B ligand (RANKL) by osteoblasts in response to interleukin (IL)-1 or IL-6 stimulation decreased by 56.8% or 50.58%, respectively, in the presence of WHI-131. WHI-131 also abrogated the formation of mature osteoclasts induced by IL-1 or IL-6 stimulation. Moreover, WHI-131 treatment decreased RANKL-induced osteoclast differentiation of bone marrow-derived macrophages, and reduced the resorbing activity of mature osteoclasts. WHI-131 further decreased the mRNA and protein expression levels of c-Fos and nuclear factor of activated T cells, cytoplasmic 1 (NFATc1) by almost twofold, and significantly downregulated the mRNA expression of the following genes: tartrate-resistant acid phosphatase (TRAP), osteoclast-associated receptor (OSCAR), DC-STAMP, OC-STAMP, ATP6v0d2, and cathepsin K (CtsK) compared with the control group. WHI-131 further suppressed the phosphorylation of protein kinase B (Akt) and degradation of inhibitor of kappa B (IκB); Ca(2+) oscillation was also affected, and phosphorylation of the C-terminal Src kinase (c-Src)-Bruton agammaglobulinemia tyrosine kinase (Btk)-phospholipase C gamma 2 (PLCγ2) (c-Src-Btk-PLCg2 calcium signaling pathway) was inhibited following WHI-131 treatment. The Janus kinase-signal transducer and activator of transcription (JAK-STAT) signaling pathway was activated by WHI-131, accompanied by phosphorylation of STAT3 Ser727 and dephosphorylation of STAT6. In osteoblasts, WHI-131 caused an approximately fourfold increase in alkaline phosphatase activity and Alizarin Red staining intensity. Treatment with WHI-131 increased the mRNA expression levels of genes related to osteoblast differentiation, and induced the phosphorylation of Akt, p38, and Smad1/5/8. Furthermore, 5-week-old ICR mice treated with WHI-131 exhibited antiresorbing effects in a lipopolysaccharide-induced calvaria bone loss model in vivo and increased bone-forming activity in a calvarial bone formation model. Therefore, the results of this study show that WHI-131 plays a dual role by inhibiting osteoclast differentiation and promoting osteoblast differentiation. Thus, WHI-131 could be a useful pharmacological agent to treat osteoporosis by promoting bone growth and inhibiting resorption.

Evaluation of a novel technique for intraperitoneal injections in mice.

Intraperitoneal injection is a common technique that safely delivers a substance into the peritoneal cavity but can induce high stress in animals. The authors have developed a new method for administering intraperitoneal injections in mice, with the goal of causing less stress during handling and injection. Here, they compare their novel technique with a conventional technique in three experiments. In the first experiment, the authors administered intraperitoneal injections of contrast medium using either technique and then used micro-computed tomography to evaluate the placement and retention of the medium. In the second and third experiments, the authors administered intraperitoneal injections or control treatments, then sampled blood to determine circulating concentrations of stress-related hormones. Imaging showed that both the novel and the conventional techniques properly delivered a contrast medium into the peritoneal cavity. The novel technique was also associated with lower concentrations of stress-related hormones than was the conventional technique. These results indicate that this novel technique might be beneficial to investigators that use intraperitoneal injections with mice.

Stauntonia hexaphylla (Lardizabalaceae) leaf methanol extract inhibits osteoclastogenesis and bone resorption activity via proteasome-mediated degradation of c-Fos protein and suppression of NFATc1 expression.

BACKGROUND: Natural plants, including common vegetables and fruits, have been recognized as essential sources for drug discovery and the development of new, safe, and economical medicaments. Stauntonia hexaphylla (Lardizabalaceae) is widely distributed in Korea, Japan, and China, and is a popular herbal supplement in Korean and Chinese folk medicine owing to its analgesic, sedative, and diuretic properties. However, the exact pharmacological effects of S. hexaphylla extract, particularly its effect on osteoclastogenesis, are not known. METHODS: Osteoclast differentiation and function were identified with tartrate-resistant acid phosphatase (TRAP) staining and bone resorption assay, and the underling mechanisms were determined by real-time RT-PCR and western blot analysis. RESULTS: S. hexaphylla was found to inhibit early-stage receptor activator of nuclear factor-κB (NF-κB) ligand (RANKL)-mediated osteoclast differentiation in bone marrow macrophages (BMMs) without cytotoxicity and bone-resorbing activity in mature osteoclasts in a dose-dependent manner. This S. hexaphylla-mediated blockade of osteoclastogenesis involved abrogation of the NF-κB, ERK, and c-Src-Btk-PLCγ2 calcium signal pathways. Interestingly, we found that S. hexaphylla down-regulated RANKL-associated c-Fos protein induction by suppressing its translation. Furthermore, ectopic overexpression of c-Fos and NFATc1 rescued the inhibition of osteoclast differentiation by S. hexaphylla. Furthermore, S. hexaphylla inhibited the c-Fos- and NFATc1-regulated expression of genes required for osteoclastogenesis, such as TRAP, OSCAR, ß3-integrin, ATP6v0d2, and CtsK. CONCLUSIONS: These findings suggest that S. hexaphylla might be useful for the development of new anti-osteoporosis agents.

CTRP3 acts as a negative regulator of osteoclastogenesis through AMPK-c-Fos-NFATc1 signaling in vitro and RANKL-induced calvarial bone destruction in vivo.

Adipokines derived from adipocytes are important factors that act as circulating regulators of bone metabolism. C1q/tumor necrosis factor (TNF)-related Protein-3 (CTRP3) is a novel adipokine with multiple effects such as lowering glucose levels, inhibiting gluconeogenesis in the liver, and increasing angiogenesis and anti-inflammation. However, the effects and the mechanisms of CTRP3 on bone metabolism, which is regulated by osteoblasts and osteoclasts, have not been investigated. Here, we found that CTRP3 inhibited osteoclast differentiation induced by osteoclastogenic factors in bone marrow cell-osteoblast co-cultures, but did not affect the ratio of receptor activator of nuclear factor κB (NF-κB) ligand (RANKL) to osteoprotegerin (OPG) induced by osteoclastogenic factors in osteoblasts. We also found that CTRP3 inhibited osteoclast differentiation from mouse bone marrow macrophages (BMMs) induced by RANKL in a dose-dependent manner without cytotoxicity. Functionally, CTRP3 inhibited the F-actin formation and bone resorbing activity of mature osteoclasts. Pretreatment with CTRP3 significantly inhibited RANKL-induced expression of c-Fos and nuclear factor of activated T-cells (NFATc1), essential transcription factors for osteoclast development. Surprisingly, the activation of AMP-activated protein kinase (AMPK) was considerably increased by pretreatment with CTRP3 for 1h. The CTRP3-stimulated AMPK activation was also maintained during RANKL-induced osteoclastogenesis. CTRP3 did not affect RANKL-induced p38, ERK, JNK, Akt, IκB, CREB, and calcium signaling (Btk and PLCγ2). These results suggest that CTRP3 plays an important role as a negative regulator of RANKL-mediated osteoclast differentiation by acting as an inhibitor of NFATc1 activation through the AMPK signaling pathway. Furthermore, CTRP3 treatment reduced RANKL-induced osteoclast formation and bone destruction in mouse calvarial bone in vivo based on micro-CT and histologic analysis. In conclusion, these findings strongly suggest that CTRP3 deserves new evaluation as a potential treatment target in various bone diseases associated with excessive osteoclast differentiation and bone destruction.

Development of a novel frontal bone defect mouse model for evaluation of osteogenesis efficiency.

The skull defect model is the existing representative osteogenesis model. The skull defect model involves monitoring osteogenesis patterns at the site of a skull defect, which has the advantages that identical defects can be induced across individual experimental animals and the results can be quantitatively evaluated. However, it can damage the cerebrum because it requires a complex surgery performed on the parietal bone. This study aims to develop a new osteogenesis model that compensates for the weak points of the existing model. Male 8-week-old imprinting control region mice were put under inhalational anesthesia, and the surgery area was disinfected with 70% ethanol prior to the creation of a 5-mm incision along the sagittal line between the glabella with a pair of scissors. The incised area was opened and, after we checked the positions of the inferior cerebral vein and the sagittal suture, a 21-gauge needle was used to make two symmetrical holes with respect to the sagittal suture 3 mm below the inferior cerebral vein and 2 mm on either side of the sagittal suture. After images were obtained using micro-computed tomography, the degree of osteogenesis was quantitatively analyzed. In addition, mRNA extracted from the site of the defect confirmed a significant increase in mRNA levels of collagen 1a, alkaline phosphatase, bone sialoprotein, osteocalcin, and Runx2, known markers for osteoblasts. The promotion of osteogenesis could be observed at the site of the defect, by histological analysis.

The Inhibitory Effect of Angelica tenuissima Water Extract on Receptor Activator of Nuclear Factor-Kappa-B Ligand-Induced Osteoclast Differentiation and Bone Resorbing Activity of Mature Osteoclasts.

Angelica tenuissima has been traditionally used in oriental medicine for its therapeutic effects in headache, toothache, and flu symptoms. It also exerts anti-inflammatory activity via the inhibition of the expression of cyclooxygenase-2 (COX-2). However, the effect of Angelica tenuissima on osteoclast differentiation has not been identified until recently. In this study, we first confirmed that Angelica tenuissima water extract (ATWE) significantly interrupted the formation of tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells (MNCs) in a dose-dependent manner without any cytotoxicity. Next, we clarified the underlying mechanisms linking the suppression effects of ATWE on the receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclastogenesis. At the molecular level, ATWE induced the dephosphorylation of c-Jun N-terminal kinase (JNK) and Akt and decreased the degradation of IκB in RANKL-dependent early signaling pathways. Subsequently, ATWE caused impaired activation of the protein and mRNA levels of c-Fos and nuclear factor of activated T cell c1 (NFATc1). Moreover, the disassembly of filamentous actin (F-actin) ring and anti-resorptive activity of mature osteoclasts were triggered by ATWE treatment. Although ATWE did not enhance osteogenesis in primary osteoblasts, our results showed that ATWE is a potential candidate for anti-resorptive agent in osteoporosis, a common metabolic bone disorder.

Ostericum koreanum Reduces LPS-Induced Bone Loss Through Inhibition of Osteoclastogenesis.

The roots of Ostericum koreanum (OK) Maximowicz have traditionally been used to produce an herbal medicine reported to possess anti-inflammatory, anti-oxidant, antimicrobial, and antitumor activities; however, its effect on bone metabolism has not yet been reported. The present study examined the effects of OK extract on lipopolysaccharide (LPS)-induced bone loss in mice by investigating bone structure and the levels of the receptor activator of nuclear factor kappa-B ligand (RANKL) and osteoprotegerin (OPG) in serum and bone marrow fluid (BMF). The effects of OK extract on osteoclastogenesis were also investigated in mouse bone marrow macrophages by examining the formation of tartrate-resistant acid phosphatase (TRAP)-positive cells, the actin ring, and bone resorption activity. OK reduced LPS-induced bone destruction in vivo via a decrease in the RANKL/OPG ratio. Furthermore, it suppressed the formation of TRAP-positive cells and the actin ring, and reduced the bone-resorbing activity of mature osteoclasts. OK also significantly down-regulated the expression of various osteoclast-specific genes. However, it did not affect osteoblast differentiation, or the expression of genes involved in this process. These results demonstrated that OK prevented LPS-induced bone loss by decreasing the RANKL/OPG ratio in serum and BMF, and inhibited osteoclast differentiation and function, suggesting that OK represents a potential therapeutic drug for the treatment of osteoclast-associated bone diseases.

Esculetin attenuates receptor activator of nuclear factor kappa-B ligand-mediated osteoclast differentiation through c-Fos/nuclear factor of activated T-cells c1 signaling pathway.

Esculetin exerts various biological effects on anti-oxidation, anti-tumors, and anti-inflammation. However, the involvement of esculetin in the bone metabolism process, particularly osteoclast differentiation has not yet been investigated. In the present study, we first confirmed the inhibitory effect of esculetin on receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast formation. We then revealed the relationship between esculetin and the expression of osteoclast-specific molecules to elucidate its underlying mechanisms. Esculetin interfered with the expression of c-Fos and nuclear factor of activated T cell c1 (NFATc1) both at the mRNA and protein level with no involvement in osteoclast-associated early signaling pathways, suppressing the expression of various transcription factors exclusively expressed in osteoclasts such as tartrate-resistant acid phosphatase (Trap), osteoclast-associated receptor (Oscar), dendritic cell-specific transmembrane protein (Dcstamp), osteoclast stimulatory transmembrane protein (Ocstamp), cathepsin K, αvß3 integrin, and calcitonin receptor (Ctr). Additionally, esculetin inhibited the formation of filamentous actin (F-actin) ring-positive osteoclasts during osteoclast differentiation. However, the development of F-actin structures and subsequent bone resorbing activity of mature osteoclasts, which are observed in osteoclast/osteoblast co-culture systems were not affected by esculetin. Taken together, our results indicate for the first time that esculetin inhibits RANKL-mediated osteoclastogenesis via direct suppression of c-Fos and NFATc1 expression and exerts an inhibitory effect on actin ring formation during osteoclastogenesis.

Dual Effect of Chrysanthemum indicum Extract to Stimulate Osteoblast Differentiation and Inhibit Osteoclast Formation and Resorption In Vitro.

The risk of bone-related diseases increases due to the imbalance between bone resorption and bone formation by osteoclasts and osteoblasts, respectively. The goal in the development of antiosteoporotic treatments is an agent that will improve bone through simultaneous osteoblast stimulation and osteoclast inhibition without undesirable side effects. To achieve this goal, numerous studies have been performed to identify novel approaches using natural oriental herbs to treat bone metabolic diseases. In the present study, we investigated the effect of Chrysanthemum indicum extract (CIE) on the differentiation of osteoclastic and osteoblastic cells. CIE inhibited the formation of TRAP-positive mature osteoclasts and of filamentous-actin rings and disrupted the bone-resorbing activity of mature osteoclasts in a dose-dependent manner. CIE strongly inhibited Akt, GSK3ß, and IκB phosphorylation in RANKL-stimulated bone marrow macrophages and did not show any effects on MAP kinases, including p38, ERK, and JNK. Interestingly, CIE also enhanced primary osteoblast differentiation via upregulation of the expression of alkaline phosphatase and the level of extracellular calcium concentrations during the early and terminal stages of differentiation, respectively. Our results revealed that CIE could have a potential therapeutic role in bone-related disorders through its dual effects on osteoclast and osteoblast differentiation.

Aconitum pseudo-laeve var. erectum inhibits receptor activator of nuclear factor kappa-B ligand-induced osteoclastogenesis via the c-Fos/nuclear factor of activated T-cells, cytoplasmic 1 signaling pathway and prevents lipopolysaccharide-induced bone loss in mice.

Aconitum pseudo-laeve var. erectum (APE) has been widely shown in herbal medicine to have a therapeutic effect on inflammatory conditions. However, there has been no evidence on whether the extract of APE is involved in the biological bone metabolism process, particularly osteoclast-mediated bone resorption. In this study, we confirmed that the administration of APE could restore normal skeletal conditions in a murine model of lipopolysaccharide (LPS)-induced bone loss via a decrease in the receptor activator of nuclear factor kappa-B ligand (RANKL)/osteoprotegerin (OPG) ratio and osteoclast number. We then investigated the effect of APE on the RANKL-induced formation and function of osteoclasts to elucidate its underlying molecular mechanisms. APE suppressed the formation of tartrate-resistant acid phosphatase (TRAP)-positive cells, as well as the bone-resorbing activity of mature osteoclasts. Furthermore, APE attenuated nuclear factor of activated T-cells, cytoplasmic 1 (NFATc1) and c-Fos without affecting any early signal pathway of osteoclastogenesis. Subsequently, APE significantly downregulated the expression of various genes exclusively expressed in osteoclasts. These results demonstrate that APE restores LPS-induced bone loss through a decrease of the serum RANKL/OPG ratio, and inhibits osteoclast differentiation and function, suggesting the promise of APE as a potential cure for various osteoclast-associated bone diseases.

Synthesis of zinc tin oxide (ZTO) nanocrystallites at room temperature through association with peptide-containing bolaamphiphile molecules.

Single crystalline zinc tin oxide (ZTO) nanocrystallites were prepared at room temperature through association with a peptide-containing bolaamphiphile molecule. The bolaamphiphile molecules self-assembled to form spherical structures with creation of ZTO nanocrystallites inside. ZTO nanocrystallite synthesis was achieved only when the bolaamphiphile molecule was present, while a mixture of amorphous Sn and Zn precipitates was formed in the absence of the bolaamphiphile molecule. The bolaamphiphile molecule is thought to stabilize the Zn(2+) and Sn(4+) precursor ions by ligation and to induce subsequent condensation forming crystalline ZTO. The ZTO formation was achieved only at a strong acidic condition that promotes dissociation and ionization of Zn and Sn precursors and represses formation of ZnO and H(2)SnO(3). The prepared ZTO nanocrystallites had almost the same band gap energy as ZTO nanoparticles prepared by the conventional hydrothermal process. The outcomes of this study indicate that the controlled mineralization of metal precursor ions in a peptide-containing bolaamphiphile molecule suspension can be an alternative method to synthesize metal oxides at room temperature, while maintaining their crystalline structure and optoelectrical properties.