Current Understanding of RANK Signaling in Osteoclast Differentiation and Maturation

박진희,이나경,이수영 한국분자세포생물학회 2017 Molecules and cells Vol.40 No.10

Osteoclasts are bone-resorbing cells that are derived from hematopoietic precursor cells and require macrophage-colony stimulating factor and receptor activator of nuclear factor-B ligand (RANKL) for their survival, proliferation, differentiation, and activation. The binding of RANKL to its receptor RANK triggers osteoclast precursors to differentiate into osteoclasts. This process depends on RANKL-RANK signaling, which is temporally regulated by various adaptor proteins and kinases. Here we summarize the current understanding of the mechanisms that regulate RANK signaling during osteoclastogenesis. In the early stage, RANK signaling is mediated by recruiting adaptor molecules such as tumor necrosis factor receptor-associated factor 6 (TRAF6), which leads to the activation of mitogen-activated protein kinases (MAPKs), and the transcription factors nuclear factor-B (NF-B) and activator protein-1 (AP-1). Activated NF-B induces the nuclear factor of activated T-cells cytoplasmic 1 (NFATc1), which is the key osteoclastogenesis regulator. In the intermediate stage of signaling, the co-stimulatory signal induces Ca2+ oscillation via activated phospholipase C2 (PLC2) together with c-Fos/AP-1, wherein Ca2+ signaling facilitates the robust production of NFATc1. In the late stage of osteoclastogenesis, NFATc1 translocates into the nucleus where it induces numerous osteoclast-specific target genes that are responsible for cell fusion and function.

Hemistepsin A ameliorates acute inflammation in macrophages via inhibition of nuclear factor-κB and activation of nuclear factor erythroid 2-related factor 2

Kim, Jae Kwang,Lee, Ji Eun,Jung, Eun Hye,Jung, Ji Yun,Jung, Dae Hwa,Ku, Sae Kwang,Cho, Il Je,Kim, Sang Chan Elsevier 2018 Food and chemical toxicology Vol.111 No.-

<P><B>Abstract</B></P> <P>Hemistepsin A (HsA) is a sesquiterpene lactone isolated from <I>Hemistepta lyrata</I> (Bunge) Bunge. We investigated the anti-inflammatory effects of HsA and sought to determine its mechanisms of action in macrophages. HsA pretreatment inhibited nitric oxide production, and reduced the expression of iNOS and COX-2 in Toll-like receptor ligand-stimulated RAW 264.7 cells. Additionally, HsA decreased the secretion of proinflammatory cytokines in lipopolysaccharide (LPS)-stimulated Kupffer cells as well as in RAW 264.7 cells. HsA inhibited phosphorylation of IKKα/β and degradation of IκBα, resulting in decreased nuclear translocation of nuclear factor-κB (NF-κB) and its transcriptional activity. Moreover, HsA phosphorylated nuclear factor erythroid 2-related factor 2 (Nrf2), increased expression levels of antioxidant genes, and attenuated LPS-stimulated H<SUB>2</SUB>O<SUB>2</SUB> production. Phosphorylation of p38 and c-Jun N-terminal kinase was required for HsA-mediated Nrf2 phosphorylation. In a D-galactosamine/LPS-induced liver injury model, HsA ameliorated D-galactosamine/LPS-induced hepatocyte degeneration and inflammatory cells infiltration. Moreover, immunohistochemical analyses using nitrotyrosine, 4-hydroxynonenal, and cleaved poly (ADP-ribose) polymerase antibodies revealed that HsA protected the liver from oxidative stress. Furthermore, HsA reduced the numbers of proinflammatory cytokine-positive cells in hepatic tissues. Thus, these results suggest HsA may be a promising natural product to manage inflammation-mediated tissue injuries through inhibition of NF-κB and activation of Nrf2.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Hemistepsin A attenuated TLRL-mediated proinflammatory response in macrophages. </LI> <LI> Hemistepsin A inhibited NF-κB, and activated Nrf2 in macrophages. </LI> <LI> Hemistepsin A ameliorated D-galactosamine/lipopolysaccharide-mediated acute liver injury in mice. </LI> </UL> </P>

Water extract of Magnolia officinalis cortex inhibits osteoclastogenesis and bone resorption by downregulation of nuclear factor of activated T cells cytoplasmic 1

심기석,김태수,하현일,이청조,이보형,김한성,박지형,마진열 한국한의학연구원 2015 Integrative Medicine Research Vol.4 No.2

Background: Magnolia officinalis cortex has been traditionally used to treat stomach and intestine diseases in traditional Korean medicine. In this study, we investigated the effect of water extract of M. officinalis cortex (WEMC) on osteoclast differentiation and function. Methods: Phytochemical characterization of WEMC was performed by high-performance liquid chromatography analysis. Osteoclast differentiation of bone marrow-derived macrophages was determined by tartrate-resistant acid phosphatase activity assay. Receptor activator of nuclear factor-κB ligand (RANKL) signaling factors and transcription factors regulating osteoclast differentiation were analyzed by Western blot and real-time polymerase chain reaction. Bone resorption function of mature osteoclasts was examined by using culture plate coated with inorganic crystalline calcium phosphate. Furthermore, the in vivo effect of WEMC on osteoporosis was examined using RANKL-induced bone loss model, characterized by micro-computed tomography and bone metabolism marker analysis. Results: WEMC inhibited RANKL-induced osteoclast differentiation and the bone resorbing activity of mature osteoclasts. WEMC contains gallic acid and honokiol as active constituents contributing to the inhibitory effect of WEMC on osteoclast differentiation. Further, WEMC suppressed RANKL-induced activation of p38 and nuclear factor-κB pathways and expression of osteoclastogenic transcription factors such as c-Fos for AP-1 and nuclear factor of activated T cells cytoplasmic 1. Ectopic overexpression of a constitutive active form of nuclear factor of activated T cells cytoplasmic 1 rescued the antiosteoclastogenic effect of WEMC. Consistent with the in vitro results, WEMC suppressed RANKL-induced trabecular bone loss in mice. Conclusion: WEMC might have a therapeutic potential to treat pathological bone diseases due to increased osteoclast differentiation and function.

Trans-10, cis-12 Conjugated Linoleic Acid Modulates Nuclear Factor-κB p65 Activity on the Production of Tumor Necrosis Factor-α in Porcine Peripheral Blood Mononuclear Cells

김영범,이일우,강지훈,양만표 한국임상수의학회 2011 한국임상수의학회지 Vol.28 No.2

Nuclear factor κB (NF-κB) is a nuclear transcription factor that modulates the expression of inflammatory cytokines such as tumor necrosis factor (TNF)-α. trans-10, cis-12 (t10c12)-conjugated linoleic acid (CLA) participates in the inhibition of TNF-α production upon lipopolysaccharide (LPS)-stimulation. However, in our previous study,t10c12-CLA enhanced the production of TNF-α by LPS-unstimulated porcine peripheral blood mononuclear cells (PBMCs) and RAW 264.7 macrophages in vitro. To resolve this apparent contradiction, we hypothesized that the effect of t10c12-CLA on TNF-α production depends on NF-κB activation induced by LPS stimulation. To test this hypothesis,we assessed the in vitro effect of t10c12-CLA on TNF-α production and NF-κB p65 activity in LPS-stimulated and LPS-unstimulated porcine PBMCs. t10c12-CLA treatment resulted in increased TNF-α production by LPS-unstimulated PBMCs but decreased TNF-α production by LPS-stimulated PBMCs. t10c12-CLA increased the degradation of inhibitory κB (IκB)-α protein and activated NF-κB p65 in LPS-unstimulated PBMCs, but had the opposite effect in LPS-stimulated PBMCs. Notably, t10c12-CLA enhanced NF-κB p65 binding activity in LPS-unstimulated PBMCs exposed to caffeic acid phenethyl ester (CAPE), a NF-κB inhibitor. Conversely, it inhibited NF-κB p65 binding activity in LPS-stimulated PBMCs exposed to CAPE. These results suggest that t10c12-CLA may have different actions under different physiological conditions, and that its effect may be associated with a change in NF-κB p65 activity.

Bavachin counteracts receptor activator of nuclear factor-κB-induced osteoclastogenesis though the suppression of nuclear factor-κB signaling pathway in RAW264.7 cells

( Bok-hee Kim ),( In-a Cho ),( Kyeong-rok Kang ),( Sook-young Lee ),( Seo-yun Jung ),( Jae-sung Kim ),( Su-gwan Kim ) 조선대학교 치의학연구원(구 조선대학교 구강생물학연구소) 2018 Oral Biology Research (Oral Biol Res) Vol.42 No.3

The aim of this study was to evaluate the biological effects and cellular signaling pathways associated with the anti-osteoclastogenesis effects of bavachin, a phytoestrogen, in the receptor activator of nuclear factor-κB ligand (RANKL)- treated RAW264.7 cells. The cell viability of RAW264.7 cells was not affected upon treatment with 5-20 μM bavachin. Furthermore, osteoclastogenesis was suppressed by bavachin in a dose-dependent manner in RAW264.7 cells treated with RANKL. Tartrate-resistant acid phosphatase, matrix metalloproteinase-9, and cathepsin K, which are closely associated with osteoclastogenesis, were significantly downregulated by bavachin in the presence of RANKL. Additionally, bavachin decreased inflammatory molecules, such as nitric oxide, inducible nitric oxide synthase, cyclooxygenase-2, and prostaglandin E2 in RAW264.7 cells treated with RANKL. Bavachin suppressed the RANKLinduced phosphorylation of nuclear factor-κB and subsequently inhibited the translocation of nuclear factor-κB from the cytosol to the nucleus. Taken together, the obtained data suggest that bavachin may prevent the osteoclast-mediated bone destructive disorders.

Inhibition of Nuclear Factor κB Activation and Cyclooxygenase-2 Expression by Aqueous Extracts of Hispanic Medicinal Herbs

Robert A. Orlando,Amanda M.,Lucy A. Hunsaker,Carolina R.,Robert E. Royer,David L. Vander Jagt,Dorothy J. Vander Jagt 한국식품영양과학회 2010 Journal of medicinal food Vol.13 No.4

Nonsteroidal anti-inflammatory drugs (NSAIDs) are a primary choice of therapy for diseases with a chronic inflammatory component. Unfortunately, long-term NSAID therapy is often accompanied by severe side effects, including cardiovascular and gastrointestinal complications. Because of this, there is critical need for identification of new and safer treatments for chronic inflammation to circumvent these side effects. Inflammatory diseases have been successfully remedied with natural herbs by many cultures. To better understand the potential of natural herbs in treating chronic inflammation and to identify their mechanism of action, we have evaluated the anti-inflammatory activities of 20 medicinal herbs commonly used in the Hispanic culture. We have established a standardized method for preparing aqueous extracts (teas) from the selected medicinal herbs and screened for inhibition of tumor necrosis factor-α-induced activation of nuclear factor κB (NF-κB), which is the central signaling pathway of the inflammatory response. A number of herbal teas were identified that exhibited significant anti-inflammatory activity. In particular, tea from the herb commonly called laurel was found to be an especially potent inhibitor of NF-κB-dependent cyclooxygenase-2 gene expression and prostaglandin E2 production in cultured murine macrophages. These findings indicate that laurel tea extract contains potent anti-inflammatory compounds that function by inhibiting the major signal transduction pathway responsible for inducing an inflammatory event. Based on these results, laurel may represent a new, safe therapeutic agent for managing chronic inflammation.

Effects of dietary Antrodia cinnamomea fermented product supplementation on metabolism pathways of antioxidant, inflammatory, and lipid metabolism pathways-a potential crosstalk

Lee M. T.,Lin W. C.,Lin L. J.,Wang S. Y.,Chang S. C.,Lee T. T. 아세아·태평양축산학회 2020 Animal Bioscience Vol.33 No.7

Objective: This study was conducted to fathom the underlying mechanisms of nutrition intervention and redox sensitive transcription factors regulated by Antrodia cinnamomea fermented product (FAC) dietary supplementation in broiler chickens. Methods: Four hundreds d-old broilers (41±0.5 g/bird) assigned to 5 groups were examined after consuming control diet, or control diet replaced with 5% wheat bran (WB), 10% WB, 5% FAC, and 10% FAC. Liver mRNA expression of antioxidant, inflammatory and lipid metabolism pathways were analyzed. Prostaglandin E2 (PGE2) concentration in each group were tested in the chicken peripheral blood mononuclear cells (cPBMCs) of 35-d old broilers to represent the stress level of the chickens. Furthermore, these cells were stimulated with 2,2′-Azobis(2-amidinopropane) dihydrochloride (AAPH) and lipopolysaccharide (LPS) to evaluate the cell stress tolerance by measuring cell viability and oxidative species. Results: Heme oxygenase-1, glutathione S-transferase, glutamate-cysteine ligase, catalytic subunit, and superoxide dismutase, and nuclear factor (erythroid-derived 2)-like 2 (Nrf2) that regulates the above antioxidant genes were all up-regulated significantly in FAC groups. Reactive oxygen species modulator protein 1 and NADPH oxygenase 1 were both rather down-regulated in 10% FAC group as comparison with two WB groups. Despite expressing higher level than control group, birds receiving diet containing FAC had significantly lower expression level in nuclear factor-kappa B (NF-κB) and other genes (inducible nitric oxide synthase, tumor necrosis factor-α, interleukin-1β, nucleotide-binding domain, leucine-rich-containing family, pyrin domain-containing-3, and cyclooxygenase 2) involving in inflammatory pathways. Additionally, except for 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase that showed relatively higher in both groups, the WB, lipoprotein lipase, Acetyl-CoA carboxylase, fatty acid synthase, fatty acid binding protein, fatty acid desaturase 2 and peroxisome proliferator-activated receptor alpha genes were expressed at higher levels in 10% FAC group. In support of above results, promoted Nrf2 and inhibited NF-κB nuclear translocation in chicken liver were found in FAC containing groups. H2O2 and NO levels induced by LPS and AAPH in cPBMCs were compromised in FAC containing diet. In 35-d-old birds, PGE2 production in cPBMCs was also suppressed by the FAC diet. Conclusion: FAC may promote Nrf2 antioxidant pathway and positively regulate lipid metabolism, both are potential inhibitor of NF-κB inflammatory pathway.

Hepatoprotective Effect of Hispidulin for the Treatment of Non-Alcoholic Fatty Liver Disease (NAFLD): Involvement of Nuclear Factor-κB and Cytochrome P450 Through Molecular Mechanism

( Dinesh Kumar Patel ),( Kanika Patel ) 대한간학회 2020 춘·추계 학술대회 (KASL) Vol.2020 No.1

Aims: Herbal drugs were mainly used for the cure and treatments of disorders until the synthetic drugs have been developed in the world. Hispidulin is a naturally occurring flavone class chemical present in Chinese herb Saussurea involucrate, Artemisia and Salvia species. Inflammations play an important role in nonalcoholic fatty liver disease (NAFLD) progression. Methods: In orders to know the importance of hispidulin against liver disorders, here in the present investigation we have analyzed all the pharmacological data of scientific research and presented in the concise form. Liver system is full of enzymes responsible for the metabolism of various molecules in the body, so effect of hispidulin on various liver enzymes have been evaluated in the present investigation through different databases analysis. Further In-silico molecular study data has been also analyzed for hispidulin against Cytochromes P450 enzyme and nuclear factor-κB to predict their molecular mechanism against nonalcoholic fatty liver disorders. Further binding energy and type of interaction of hispidulin against Cytochromes P450 and nuclear factor-κB were also analyzed through data analysis of various scientific research in the present investigation to predict their molecular mechanism. Results: From the analysis of the scientific data of different research, we found that hispidulin interacted liver mitochondria and inhibited enzymatic activities, however hispidulin also counteract reduced glutathione depletion induced by bromobenzene in starved mice. Scientific investigation revealed the importance of hispidulin against nonalcoholic fatty liver disease through different data analysis as hispidulin interact liver enzymes and regulated them significantly. Molecular study data analysis revealed the interaction of hispidulin and Cytochromes P450 enzyme and nuclear factor-κB as it showed negative binding energy and interaction with the respective ligand molecule. From the present investigation it was found that hispidulin have hepatoprotective effects in the nonalcoholic fatty liver disease. Conclusions: Present work summarized pharmacological importance of hispidulin against nonalcoholic fatty liver disease and this work will be beneficial to the researcher for the development of novel molecule against hepatic disorders.

RAW 264.7 세포에서 음양곽(淫羊藿) 물 추출물의 nuclear factor-κB 억제를 통한 항염증 효과

정지윤 ( Ji Yun Jung ),변성희 ( Sung Hui Byun ),박정아 ( Chung A Park ),조일제 ( Il Je Cho ),김상찬 ( Sang Chan Kim ) 대한본초학회 2018 大韓本草學會誌 Vol.33 No.2

Objectives : Epimedii Herba has been frequently used in Korean Traditional Medicine to treat impotence, spermatorrhoea, exophthalmos, and forgetfulness. Present study investigated anti-inflammatory effects of Epimedii Herba water extract (EWE) and attempted to elucidate molecular mechanisms involved. Methods : To explore anti-inflammatory effects of EWE, RAW 264.7 cells, a murine macrophage cell line, were pretreated with 10-100 ㎍/㎖ of EWE, and then subsequently exposed to 1 ㎍/㎖ of lipopolysaccharide (LPS). Levels of nitric oxide (NO), interleukin-6, interleukin-1β, and tumor necrosis factor-α were monitored in the medium. Expression levels of inducible nitric oxide synthase and cyclooxygenase-2 were determined by immunoblot and real-time PCR analyses. Signaling pathways related with nuclear factor-κB (NF-κB) and mitogen-activated protein kinases were monitored to elucidate molecular mechanisms involved. Finally, the role of three flavonoid compounds in EWE on LPS-mediated NO production were investigated. Results : In conditioned medium, pretreatment of EWE (100 ㎍/㎖) significantly inhibited LPS-stimulated NO and pro-inflammatory cytokine production. In addition, EWE attenuated the expressions of inducible nitric oxide synthase and cyclooxygenase-2 by LPS. EWE prevented the phosphorylation and degradation of inhibitory κBα, nuclear translocation of NF-κB, and DNA binding of NF-κB, while EWE did not change the phosphorylation of mitogenactivated protein kinases by LPS. Moreover, icariin, icaritin, and quercetin partly, but significantly, inhibited the LPS-stimulated NO production. Conclusions : These results suggest that EWE has an ability to prevent inflammation in macrophages through inhibition of NF-κB signaling pathway.

Korean Red Ginseng mitigates spinal demyelination in a model of acute multiple sclerosis by downregulating p38 mitogen-activated protein kinase and nuclear factor-kB signaling pathways

이민정,장병준,오세관,나승열,조익현 고려인삼학회 2018 Journal of Ginseng Research Vol.42 No.4

Background: The potential therapeutic values of Korean Red Ginseng extract (KRGE) in autoimmune disorders of nervous system have not been fully investigated. Methods: We used an acute experimental autoimmune encephalomyelitis animal model of multiple sclerosis and determined the effects and mechanism of KRGE on spinal myelination. Results: Pretreatment with KRGE (100 mg/kg, orally) for 10 days before immunization with myelin basic protein (MBP)68e82 peptide exerted a protective effect against demyelination in the spinal cord, with inhibited recruitment and activation of immune cells including microglia, decreased mRNA expression of detrimental inflammatory mediators (interleukin-6, interferon-g, and cyclooxygenase-2), but increased mRNA expression of protective inflammatory mediators (insulin-like growth factor b1, transforming growth factor b, and vascular endothelial growth factor-1). These results were associated with significant downregulation of p38 mitogen-activated protein kinase and nuclear factor-kB signaling pathways in microglia/macrophages, T cells, and astrocytes. Conclusion: Our findings suggest that KRGE alleviates spinal demyelination in acute experimental autoimmune encephalomyelitis through inhibiting the activation of the p38 mitogen-activated protein kinase/nuclear factor-kB signaling pathway. Therefore, KRGE might be used as a new therapeutic for autoimmune disorders such as multiple sclerosis, although further investigation is needed.