Regulatory role of the transcriptional coactivator Mediator 1 in skin hair regeneration and its mechanisms / 中华皮肤科杂志

Objective:To investigate the effect of the transcriptional coactivator Mediator 1 (Med1) on mouse hair regeneration, and to explore potential mechanisms.Methods:Med1 flox/flox C57BL/6J mice were mated with K14-Cre mice, and the mice with epidermis-specific knockout of Med1 gene, namely K14-Cre-expressing Med1 flox/flox mice (knockout group) , were obtained by using the Cre-Loxp system, while Med1 flox/flox mice without K14-Cre expression served as control group. Mice in the two groups (3 mice in each group) were raised together for 8 weeks followed by dorsal hair removal. Hair regeneration was observed for 12 consecutive days after hair removal. After 12 days, all mice in the two groups were sacrificed, their depilated and non-depilated dorsal skin tissues were resected, and total RNA was extracted from the tissues. Real-time quantitative PCR was performed to determine the mRNA expression of hair keratin genes, vitamin D receptor/β-catenin pathway-related genes, and genes associated with maintenance of hair follicle stem cell proliferation and quiescence. Paraffin-embedded sections of depilated and non-depilated mouse skin tissues were prepared, and immunofluorescence staining was conducted to determine the number of stem cells in the hair follicle bulge. Two-independent-sample t test was used for comparisons between two groups. Results:From days 0 to 12 after depilation, hair regeneration was delayed in the depilated skin area in the knockout group compared with the control group. Real-time quantitative PCR showed significantly decreased mRNA relative expression levels of hair keratin genes Ha1 and Krt2-16, vitamin D receptor/β-catenin pathway-related genes S100a3, Dlx3 and Tubb3, and genes associated with maintenance of hair follicle stem cell proliferation and quiescence including Lhx2, Sox9 and Nfatc1 in the depilated skin tissues in the knockout group (22.09 ± 12.32, 2.07 ± 0.20, 0.02 ± 0.01, 12.36 ± 2.12, 1.75 ± 0.46, 0.39 ± 0.02, 4.42 ± 0.76, 0.44 ± 0.07, respectively) compared with the control group (70.53 ± 9.46, 7.76 ± 0.49, 0.05 ± 0.01, 26.16 ± 2.96, 2.60 ± 0.14, 0.71 ± 0.09, 11.93 ± 0.42, 0.75 ± 0.04, respectively; t = 5.40, 18.64, 3.89, 6.57, 3.04, 6.10, 15.03, 6.18, respectively, all P < 0.05) . Immunofluorescence staining showed that the number of CD34 +K15 + hair follicle stem cells in the hair follicle bulge in both depilated and non-depilated skin tissues was significantly lower in the knockout group than in the control group. Conclusion:Med1 gene knockout may down-regulate the expression of downstream genes of the vitamin D receptor/β-catenin pathway and genes associated with maintenance of hair follicle stem cell proliferation and quiescence (Sox9, Nfatc1 and Lhx2) , and reduce the number of hair follicle stem cells, leading to hair follicle differentiation disorder and hair regeneration delay.

Regulation of signaling pathways associated with cardiac valve development / 国际儿科学杂志

Congenital heart disease is the most common birth defect in China,of which heart valve dysplasia is an important phenotype.Heart valve development is an important process of embryonic development,which is regulated by a variety of signaling pathways.If the process of proliferation,differentiation or migration of endothelial cells and cardiomyocytes is abnormal,the heart valve will develop abnormally and the valvular heart disease may occur.Tissue explant systems and various animal model experiments have demonstrated that multiple signaling pathways interact to form a vast regulatory network that collectively regulates the development of heart valves.This review will highlight the nost intensively studied signaling pathways in epithelial-mesenchymal transition,including VEGF,NFATc1,Notch,Wnt,TGF-β,ErbB,and NF1 signaling pathways.

Inhibitory Effect of Rosae Multiflorae Fructus Extracts on the Receptor Activator of NF-κB Ligand-Induced Osteoclastogenesis through Modulation of P38- and Ca2⁺-Mediated Nuclear Factor of Activated T-Cells Cytoplasmic 1 Expression

BACKGROUND: Rosae Multiflorae fructus (RMF), known to have anti-inflammatory and antioxidant properties, has been used as a traditional remedy for inflammatory diseases such as arthritis in Eastern Asia. However, its effect on osteoclasts, which play a crucial role in resorptive inflammatory bone diseases, is yet to be elucidated.METHODS: The effect of extract of RMF (RMF-E) on receptor activator of nuclear factor-κB ligand (RANKL)-mediated osteoclastogenesis was examined by tartrate-resistant acid phosphatase (TRAP) staining, real-time polymerase chain reaction and western blot analysis. In addition, RANKL-induced Ca2⁺-oscillation was also investigated.RESULTS: RMF-E remarkably inhibited TRAP+-osteoclast and resorptive pit formation in a dose-dependent manner. In addition, the expression of c-Fos and nuclear factor of activated T-cells cytoplasmic, known as pivotal transcription factors for osteoclast formation in vitro and in vivo, and that of the osteoclast differentiation markers such as Acp5, Oscar, CtsK, Atp6v0d2, Tm7sf4, and Nfatc1 were significantly decreased by RMF-E treatment during osteoclastogenesis. The inhibitory effect of RMF-E on RANKL-induced osteoclastogenesis was caused by the suppression of p38 mitogen-activated protein kinase activation, and RANKL-induced Ca2⁺-oscillation removal via inactivation of Bruton's tyrosine kinase (BTK), and subsequently phospholipase C-γ2.CONCLUSIONS: RMF-E negatively regulates osteoclast differentiation and formation. These findings suggest the possibility of RMF-E as a traditional therapeutic agent against osteoclast-related bone disorders such as osteoporosis, rheumatoid arthritis, and periodontitis.

Regulation of signaling pathways associated with cardiac valve development / 国际儿科学杂志

Congenital heart disease is the most common birth defect in China, of which heart valve dysplasia is an important phenotype.Heart valve development is an important process of embryonic development, which is regulated by a variety of signaling pathways.If the process of proliferation, differentiation or migration of endothelial cells and cardiomyocytes is abnormal, the heart valve will develop abnormally and the valvular heart disease may occur.Tissue explant systems and various animal model experiments have demonstrated that multiple signaling pathways interact to form a vast regulatory network that collectively regulates the development of heart valves.This review will highlight the nost intensively studied signaling pathways in epithelial-mesenchymal transition, including VEGF, NFATc1, Notch, Wnt, TGF-β, ErbB, and NF1 signaling pathways.

Simvastatin promotes murine osteoclasts apoptosis through NFATc1 pathway / 南方医科大学学报

OBJECTIVE@#To explore the mechanism by which simvastatin (SIM) regulates osteoclast apoptosis.@*METHODS@#Murine macrophage RAW264.7 cells were divided into 5 groups, namely group A (control group), group B (sRANKL+ M-CSF), group C (SIM+sRANKL+M-CSF), group D (VIVIT peptide+sRANKL+ M-CSF), and group E (SIM+VIVIT peptide+sRANKL+M-CSF). WST-1 assay was used to assess the effects of simvastatin on the proliferation activity of the osteoclasts, and flow cytometry was performed to analyze the effects of SIM and VIVIVIT peptide (a NFATc1 pathway inhibitor) on apoptosis of the osteoclasts. The translocation of NFATc1 into the nucleus was investigated using immunofluorescence assay, and Western blotting was employed to assess the effect of SIM on the phosphorylation of NFATc1 in the nucleus.@*RESULTS@#WST-1 assay showed that SIM (1×10 mol/L) treatment for 24 and 48 h significantly inhibited the proliferation of the osteoclasts (=0.039 and 0.022, respectively). Compared with the control group, the SIM-treated osteoclasts exhibited significantly reduced cell percentage in G0/G1 phase (=0.041) and increased cells in sub-G1 phase (=0.028) with obvious cell apoptosis. DAPI staining and flow cytometry showed that both SIM and VIVIVIT peptide alone significantly promoted osteoclast apoptosis (=0.002 and 0.015, respectively), and their combination produced a similar pro-apoptosis effect (=0.08). Immunofluorescence and Western blotting showed that SIM significantly inhibited the intranuclear translocation of NFATc1 and the phosphorylation of NFATc1 pathway protein (=0.013).@*CONCLUSIONS@#SIM promotes osteoclast apoptosis through NFATc1 signaling pathway.

Inhibition of Osteoclast differentiation based on precipitation time of titanium surfaces immersed in modified simulated body fluid / 대한치과보철학회지

PURPOSE: The purpose of this study is to investigate the changes of osteoclast differentiation inhibition according to the period of precipitation when titanium disks were immersed in Modified simulated body fluid (mSBF). MATERIALS AND METHODS: Titanium alloy (Ti grade III) disks with machined surfaces and anodized surfaces were immersed in distilled water and mSBF, respectively. The immersion periods were 7 days, 14 days, 21 days and 28 days, and the control group was immersed in distilled water for each period. RAW 264.7 cells capable of differentiating into osteoclasts were used to measure the number of adherent cells, the measurement of TRAP activity, and the expression pattern of NFATc1 by western blotting. RESULTS: The degree of inhibition of osteoclast differentiation was found to be statistically significant when the disks were immersed in mSBF for more than 14 days on both machined surfaces and anodized surfaces. There was no correlation between immersion time and cell attachment. When the disks were immersed for more than 14 days, TRAP activity was decreased and NFATc1 expression was inhibited. Futhermore, the decrease in TRAP activity and the inhibition of NFATc1 expression remained unchanged. CONCLUSION: Immersion of titanium disks in mSBF for more than 14 days can prevent RAW 264.7 cells from differentiating into osteoclasts. Inhibition activity does not change even if the immersion period is for more than 14 days.

IL-38 inhibits osteoporosis via regulating PI3K/Akt/GSK3β/NFATc1 signaling pathway / 中国免疫学杂志

Objective:To investigate the role and mechanism of IL-38 in inhibiting osteoporosis.Methods:A total of 138 cases of patients with osteoporosis in our hospital from June 2014 to December 2016 were recruited.Another 120 cases of fracture surgery patients without osteoporosis were selected as control.Serum levels of IL-38 in different groups were determined using a commercially available sandwich ELISA (Enzyme-Linked ImmunoSorbent Assay).Construction of IL-38-C57BL/6J transgenic mice,the wild type and IL-38 transgenic mice were set to sham operation group (Sham),operation group (ovariectomy,group OVX) respectively.After 8 weeks of the operation,the serum level of alkaline phosphatase(ALP),calcium and phosphorus were detected.The bilateral femur and spine of mice were collected after sacrifice,the morphology and structure of the femur were analyzed,and the bone density was measured by bone density meter.The bone marrow stromal cells(BMSCs) were isolated and the invitro proliferation ability of BMSCs were meas-ured.Western blot were used to detect the phosphorylation level of PI3K,Akt,GSK3β and NFATc1 in BMSCs.After transfection of IL-38 into mouse osteoblast MC3T3-E1 cell,the phosphorylation level of PI3K,Akt,GSK3β and NFATc1 were detected by Western blot.Apoptosis of MC3T3-E1 cells were detected by flow cytometry.Results:The serum level of IL-38 in patients with osteoporosis were significant lower than control group(P<0.05).The serum level of estrogen,calcium and phosphorus in OVX group of wild type and IL-38 transgenic mice were significant lower than the sham operation group(P<0.05),while the level of ALP was significant higher than sham operation group (P<0.05),but the serum level of calcium and phosphorus in OVX group of IL-38 transgenic mice were significant higher than wild type mice(P<0.05).The pathological section of femur and spine BMD showed that the bone tissue in wild type mice and IL-38 transgenic mice in OVX group were damaged and the bone density decreased significantly,but IL-38 transgenic mice was significant better than wild type mice (P<0.05).The proliferation ability of BMSCs in OVX group of IL-38 transgenic mice was significant higher than wild type mice (P<0.05).The phosphorylation level of PI3K,Akt and NFATc1 in OVX group of IL-38 transgenic mice were significant lower than wild type mice(P<0.05),while the phosphorylation level of GSK3β was significant higher than wild type mice (P<0.05).After transfection of IL-38 into MC3T3-E1 cell,the phosphorylation level of PI3K,Akt and NFATc1 were significant decreased (P<0.05),while the phosphorylation level of GSK3β was significant increased (P<0.05).Flow cytometry assay showed that IL-38 transfection significant decreased the apoptosis of MC3T3-E1 cells(P<0.05).Conclusion:The serum level of IL-38 in patients with osteoporosis is decreased significantly.IL-38 may inhibit the proliferation of BMSCs and inhibit the apoptosis of osteoblasts by regulating the PI3K/Akt/GSK3β/NFATc1 signaling pathway.

Curcumin Reduces the Noise-Exposed Cochlear Fibroblasts Apoptosis

Abstract Introduction The structural changes underlying permanent noise-induced hearing loss (NIHL) include loss of the sensory hair cells, damage to their stereocilia, and supporting tissues within the cochlear lateral wall. Objective The objective of this study is to demonstrate curcumin as a safe and effective therapeutic agent in the prevention and treatment for fibroblasts damage within the cochlear supporting tissues and lateral wall through cell death pathway. Methods We divided 24 Rattus norvegicus into 4 groups, Group 1: control; Group 2: noise (þ); Group 3: noise (þ), 50 mg/day curcumin (þ); Group 4: noise (þ), 100 mg/day curcumin (þ). We provided the noise exposure dose at 100 dB SPL for two hours over two weeks and administered the curcumin orally over two weeks. We examined all samples for the expressions of calcineurin, nuclear factor of activated T-cells cytoplasmic 1 (NFATc1), and apoptotic index of cochlear fibroblasts. Results We found significant differences for the expressions of calcineurin (p< 0.05) in all groups, significant differences for the expressions of NFATc1 (p< 0.05) in all groups, except in Groups 1 and 4, and significant differences for the apoptotic index (p< 0.05) in all groups. Conclusion Curcumin proved to be potentially effective in the prevention and treatment for fibroblasts damage within the cochlear supporting tissues and lateral wall regarding the decreased expression of calcineurin, NFATc1, and apoptotic index of cochlear fibroblasts.

NFATc1 promotes vascular generation of epithelial ovarian cancer trans-planted tumor by regulating CXCR2, FGF-2 and PDGF-BB / 中国病理生理杂志

AIM:To investigate the role of NFATc1 in vascular generation in the nude mice transplanted with human ovarian cancer SKOV3 cells.METHODS: NFATc1 expression was silenced by siRNA in SKOV3 cells.Human ovarian cancer transplantation nude mouse model was established by transplanting with SKOV3 cells in which the NFATc1 gene was silenced by siRNA technique.The expression of NFATc1, CXCR2, FGF-2 and PDGF-BB at mRNA and protein levels was determined by RT-PCR, Western blotting and immunohistochemical staining.The tumor growth, angiogenesis and lymphangiogenesis were also observed.RESULTS:Over-expression of NFATc1 was observed in human ovarian cancer tissues.The silencing of NFATc1 expression by siRNA decreased tumorigenesis of transplanted ovarian cancer cells in the nude mice, reduced tumor vascular generation and inhibited the expression of CXCR2, FGF-2 and PDGF-BB at mRNA and protein levels.CONCLUSION:NFATc1 is overexpressed in ovarian cancer.NFATc1 silencing regulates the tumor vascu-lar generation.NFATc1 thus has potential as a therapeutic target and for use in the diagnosis and evaluating prognosis of epithelial ovarian cancer.

Attenuation of RANKL-induced Osteoclast Formation via p38-mediated NFATc1 Signaling Pathways by Extract of Euphorbia Lathyris L

BACKGROUND: Osteoclasts are the only cell type capable of breaking down bone matrix, and its excessive activation is responsible for the development of bone-destructive diseases. Euphorbia lathyris L. (ELL) is an herbal plant that belongs to the Euphorbiaceae family. This study investigated the effects of the methanol extract of the aerial part of ELL on receptor activator of nuclear factor-kappa B ligand (RANKL)-induced osteoclast formation and signaling pathways. METHODS: Osteoclasts were formed by co-culturing mouse bone marrow with osteoblasts or by culturing mouse bone marrow-derived macrophages (BMMs) with macrophage colony-stimulating factor (M-CSF) and RANKL. Bone resorption assays were performed using dentine slices. The expression level of mRNA was analyzed by real-time polymerase chain reaction (PCR) or reverse transcription (RT)-PCR. Western blotting assays were performed to detect the expression or activation level of proteins. RESULTS: ELL inhibited RANKL-induced osteoclast formation without cytotoxicity. Furthermore, the RANKL-stimulated bone resorption was diminished by ELL. Mechanistically, ELL blocked the RANKL-triggered p38 mitogen-activated protein kinase (MAPK) phosphorylation, which resulted in the suppression of the expression of c-Fos and nuclear factor of activated T cells (NFATc1). In osteoblasts, ELL had little effect on the mRNA expression of RANKL and osteoprotegerin (OPG). CONCLUSIONS: The present data suggest that ELL has an inhibitory effect on osteoclast differentiation and function via downregulation of the p38/c-Fos/NFATc1 signaling pathways. Thus, ELL could be useful for the treatment of bone diseases associated with excessive bone resorption.

Protective effect of cinnamic aldehyde on hormone-induced osteoclasts differentiation and its molecular mechanisms / 中国药理学通报

Aim To investigate the protective effect of cinnamic aldehyde ( CA ) on hormone-induced osteo-clasts proliferation and bone resorption in vitro and its molecular mechanisms. Methods RAW264. 7 cells induced into osteoclast were treated with RANKL and M-CSF and then were divided into control group, dexa-methasone ( DEX ) group and different doses of CA (11. 6, 23. 2, 46. 4 μg·L-1 ) groups. OCs were ob-served after tartrate resistant acid phosphatase( TRAP) staining. The cell proliferation was determined by MTT assay at different time points. The expression levels of TRACP5 b in cell cultured supernatants were measured by ELISA. RT-PCR technique was applied to examine the transcriptional levels of RANK and NFATc1 . Re-sults In MTT assay, the proliferation of osteoclasts stimulated by dexamethasone was promoted seriously compared with negative control group ( P < 0. 05 ) . Meanwhile, DEX could strengthen the content of TRACP5 b and up-regulate the expressions of RANK and NFATc1 mRNA. After administration of CA, the proliferation was inhibited, while the enhanced expres-sion of TRAP5b was reversed,and the over-expressions of RANK and NFATc1 mRNA were obviously down-regulated in a time-and-dose-dependent manner ( P <0. 05 ) . Conclusion The results suggest that CA in-hibits proliferation and bone resorption of osteoclast in-duced by DEX, which may be mediated by down-regu-lation of RANK and NFATc1 mRNA.

MSK1 regulates RANKL-induced NFATc1 expression through CREB and c-Fos / 동물의과학연구지

Osteoclasts originated from hematopoietic stem cells are multi-nucleated cells that can resorb the bone matrix. Receptor activator of nuclear factor kappa-B (RANK)/RANK ligand (RANKL) signaling pathway is crucial for the differentiation and activation of osteoclasts. In this study, we investigated for the first time whether or not RANKL induced mitogen- and stress-activated kinase 1 (MSK1) phosphorylation at Ser 376. Activation of MSK1 was detected as soon as 5 min after RANKL stimulation and sparsely detected at 30 min after stimulation. RANKL-induced MSK1 phosphorylation occurred in a dose-dependent manner. MSK1 is known as a downstream signaling molecule of cAMP-dependent protein kinase (PKA). Treatment with the PKA inhibitor H89 significantly suppressed c-Fos and nuclear factor of activated T-cells, cytoplasmic 1 (NFATc1) induction upon RANKL stimulation. In addition, cAMP response element-binding protein (CREB) phosphorylation was extremely inhibited by H89 treatment. Mitogen-activated protein kinases (MAPKs) have been investigated for induction of MSK1 phosphorylation. Specific signaling pathway inhibitors for p38 and extracellular signal-regulated kinases (ERKs) significantly blocked RANKL-induced MSK1 activation. Finally, as a downstream effector of the p38-MSK1 pathway, c-Fos transcriptional activity was determined. RANKL-mediated elevation of c-Fos transcriptional activity was significantly suppressed by p38 inhibitor. Moreover, a dominant negative form of CREB suppressed activation of NFATc1. In conclusion, RANKL-stimulated MSK1 phosphorylation could play a role in induction of NFATc1 through CREB and c-Fos activation as a downstream molecule of p38, ERK MAPKs, and PKA. Our results support basic information for the development of osteoclast specific inhibitors.

Cyclosporine Stimulates the Expression of CDK4 through NFATc1 Suppression: Anagen Phase Hair Induction / 대한피부과학회지

BACKGROUND: Hair growth is spontaneously activated from quiescent bulge stem cells or is activated from precocious anagen. Upon spontaneous activation of hair growth or activation induced by nuclear factor of activated T cells c1 (NFATc1) inhibitors, NFATc1 expression is lost and cyclin dependent kinase (CDK4) repression is relieved. OBJECTIVE: The purpose of this study was to investigate the mechanisms of cyclosporine as a hair cycle regulator in the treatment of Alopecia areata (AA). METHODS: In this study, we planned to investigate the hair growing properties of cyclosporine in vitro conditions. Briefly, the effects of different concentrations of cyclosporine (200, 500, 1,000, 2,000 mmol) on the growth of cultured hair follicles were examined through the expression of NFATc1 and CDK4. RESULTS: NFATc1 was downregulated and CDK4 expression was upregulated especially in the bulge areas, outer root sheath and hair bulb matrix cells as the concentration of cyclosporine increased. CONCLUSION: Cyclosporine induces CDK4 expression by NFATc1 suppression, which acts to relieve repressed CDK4, resulting in hair growth. In conclusion, cyclosporine is one of the candidates as a therapeutic agent for the treatment of hair loss.

NFATc1 and NFATc3 is Involved in the Expression of Receptor Activator of NF-kappaB Ligand in Activated T Lymphocytes

Receptor activator of NF-kappaB ligand (RANKL) is an essential cytokine for osteoclast differentiation, activation and survival. T lymphocytes such as T17 cells, a subset of T helper cells that produce IL-17, play an important role in rheumatoid arthritic bone resorption by producing inflammatory cytokines and RANKL. It has not yet been clearly elucidated how T cell activation induces RANKL expression. T cell receptor activation induces the activation of nuclear factor of activated T cell (NFAT) and expression of its target genes. In this study, we examined the role of NFAT in T cell activation-induced RANKL expression. EL-4, a murine T lymphocytic cell line, was used. When T cell activation was induced by phorbol 12-myristate 13-acetate (PMA) and ionomycin, RANKL expression increased in a time-dependent manner. In the presence of cyclosporin, an inhibitor of NFAT activation, this PMA/ionomycin-induced RANKL expression was blocked. Overexpression of either NFATc1 or NFATc3 induced RANKL expression. Chromatin immunoprecipitation results demonstrated that PMA/ionomycin treatment induced the binding of NFATc1 and NFATc3 to the mouse RANKL gene promoter. These results suggest that NFATc1 and NFATc3 mediates T cell receptor activation-induced RANKL expression in T lymphocytes.

The Molecular Mechanism of Baicalin on RANKL-induced Osteoclastogenesis in RAW264.7 Cells

This study examined the anti-osteoclastogenic effects of baicalin on receptor activator of NF-kB ligand (RANKL)-induced RAW264.7 cells. Baicalin is a flavonoid that is produced by Scutellaria baicalensis and is known to have multiple biological properties, including antibacterial, anti-inflammatory and analgesic effects. The effects of baicalin on osteoclasts were examined by measuring 1) cell viability; 2) the formation of tartrate-resistant acid phosphatase (TRAP) (+) multinucleated cells; 3) RANK/RANKL signaling pathways and 4) mRNA levels of osteoclast-associated genes. Baicalin inhibited the formation of RANKL-stimulated TRAP (+) multinucleated cells and also suppressed the RANKL-stimulated activation of p-38, ERK, cSrc and AKT signaling. Baicalin also inhibited the RANKL-stimulated degradation of IkappaB in RAW264.7 cells. In addition, the RANKL-stimulated induction of NFATc1 transcription factors was found to be abrogated by this flavonoid. Baicalin was further found to decrease the mRNA expression of osteoclast-associated genes, including carbonic anhydrase II, TRAP and cathepsin K in the RAW264.7 cells. Our data thus demonstrate that baicalin inhibits osteoclastogenesis by inhibiting the RANKL-induced activation of signaling molecules and transcription factors in osteoclast precursors.

Amorphigenin inhibits Osteoclast differentiation by suppressing c-Fos and nuclear factor of activated T cells / 대한해부학회지

Among the several rotenoids, amorphigenin is isolated from the leaves of Amopha Fruticosa and it is known that has anti-proliferative effects and anti-cnacer effects in many cell types. The main aim of this study was to investigate the effects of amorphigenin on osteoclast differentiation in vitro and on LPS treated inflammatory bone loss model in vivo. We show here that amorphigenin inhibited RANKL-induced osteoclast differentiation from bone marrow macrophages in a dose dependent manner without cellular toxicity. Anti-osteoclastogenic properties of amorphigenin were based on a down-regulation of c-fos and NFATc1. Amorphigenin markedly inhibited RANKL-induced p38 and NF-kappaB pathways, but other pathways were not affected. Micro-CT analysis of the femurs showed that amorphigenin protected the LPS-induced bone loss. We concluded that amorphigenin can prevent inflammation-induced bone loss. Thus we expect that amorphigenin could be a treatment option for bone erosion caused by inflammation.