Targeting the TNF-α-TNFR interaction with EGCG to block NF-κB signaling in human synovial fibroblasts.

The tumor necrosis factor alpha (TNF-α)-TNF-α receptor (TNFR) interaction plays a central role in the pathogenesis of various autoimmune diseases, particularly rheumatoid arthritis, and is therefore considered a key target for drug discovery. However, natural compounds that can specifically block the TNF-α-TNFR interaction are rarely reported. (-)-Epigallocatechin-3-gallate (EGCG) is the most active, abundant, and thoroughly investigated polyphenolic compound in green tea. However, the molecular mechanism by which EGCG ameliorates autoimmune arthritis remains to be elucidated. In the present study, we found that EGCG can directly bind to TNF-α, TNFR1, and TNFR2 with similar µM affinity and disrupt the interactions between TNF-α and TNFR1 and TNFR2, which inhibits TNF-α-induced L929 cell death, blocks TNF-α-induced NF-κB activation in 293-TNF-α response cell line, and eventually leads to inhibition of TNF-α-induced NF-κB signaling pathway in HFLS and MH7A cells. Thus, regular consumption of EGCG in green tea may represent a potential therapeutic agent for the treatment of TNF-α-associated diseases.

ATP supplementation suppresses UVB-induced photoaging in HaCaT cells via upregulation of expression of SIRT3 and SOD2.

BACKGROUND: Skin photoaging is the damage caused by excessive exposure to ultraviolet (UV) irradiation. We investigated the effect of adenosine triphosphate (ATP) supplementation on UVB-induced photoaging in HaCaT cells and its potential molecular mechanism. MATERIALS AND METHODS: The toxicity of ATP on HaCaT cells was examined by the MTT assay. The effects of ATP supplementation on the viability and apoptosis of HaCaT cells were determined by crystal-violet staining and flow cytometry, respectively. Cellular and mitochondrial ROS were stained using fluorescent dyes. Expression of Bax, B-cell lymphoma (Bcl)-2, sirtuin (SIRT)3, and superoxide dismutase (SOD)2 was measured via western blotting. RESULTS: ATP (1, 2 mM) exerted no toxic effect on the normal growth of HaCaT cells. UVB irradiation caused the apoptosis of HaCaT cells, and ATP supplementation inhibited the apoptosis induced by UVB significantly, as verified by expression of Bax and Bcl-2. UVB exposure resulted in accumulation of cellular and mitochondrial reactive oxygen species (ROS), but ATP supplementation suppressed these increases. Expression of SIRT3 and SOD2 was decreased upon exposure to UVB irradiation but, under ATP supplementation, expression of SIRT3 and SOD2 was reversed, which was consistent with the reduction in ROS level observed in ATP-treated HaCaT cells after exposure to UVB irradiation. CONCLUSIONS: ATP supplementation can suppress UVB irradiation-induced photoaging in HaCaT cells via upregulation of expression of SIRT3 and SOD2.

Tereticornate A suppresses RANKL-induced osteoclastogenesis via the downregulation of c-Src and TRAF6 and the inhibition of RANK signaling pathways.

Excessive osteoclast differentiation and activation are closely associated with the development and progression of osteoporosis. Natural plant-derived compounds that can inhibit osteoclastogenesis are an efficient strategy for the prevention and treatment of osteoporosis. Tereticornate A (TA) is a natural terpene ester compound extracted from the leaves and branches of Eucalyptus gracilis, with antiviral, antibacterial, and anti-inflammatory activities. However, the effect of TA on osteoclastogenesis and the underlying molecular mechanism remain unclear. Based on the key role of the NF-κB pathway in the regulation of osteoclastogenesis and the observation that TA exhibits an anti-inflammatory effect by inhibiting NF-κB activity, we speculated that TA could exert anti-osteoclastogenesis activity. Herein, TA could inhibit the RANKL-induced osteoclast differentiation and formation of F-actin rings in RAW 264.7 cells. Mechanistically, TA downregulated the expression of c-Src and TRAF6, and also suppressed the RANKL-stimulated canonical RANK signaling pathways, including AKT, MAPK (p38, JNK, and ERK), and NF-κB; ultimately, downregulating the expression of NFATc1 and c-Fos, the key transcriptional factors required for the expression of genes (e.g., TRAP, cathepsin K, ß-Integrin, MMP-9, ATP6V0D2, and DC-STAMP) that govern osteoclastogenesis. Our findings demonstrated that TA could effectively inhibit RANKL-induced osteoclastogenesis via the downregulation of c-Src and TRAF6 and the inhibition of RANK signaling pathways. Thus, TA could serve as a novel osteoclastogenesis inhibitor and might have beneficial effects on bone health.

Niloticin inhibits osteoclastogenesis by blocking RANKL-RANK interaction and suppressing the AKT, MAPK, and NF-κB signaling pathways.

Dysregulation of osteoclasts or excessive osteoclastogenesis significantly -contributes to the occurrence and development of osteolytic diseases, including osteoporosis, inflammatory bone erosion, and tumor-induced osteolysis. The protein-protein interaction between the receptor activator of nuclear factor (NF)-κB (RANK) and its ligand (RANKL) mediates the differentiation and activation of osteoclasts, making it a key therapeutic target for osteoclastogenesis inhibition. However, very few natural compounds exerting anti-osteoclastogenesis activity by inhibiting the RANKL-RANK interaction have been found. Niloticin is a natural tetracyclic triterpenoid compound with anti-viral, antioxidative, and mosquitocidal activities. However, its role in osteoclastogenesis remains unknown. The present study found that niloticin directly binds to RANK with an equilibrium dissociation constant of 5.8 µM, blocking RANKL-RANK interaction, thereby inhibiting RANKL-induced AKT, MAPK (p38, JNK, and ERK1/2), and NF-κB (IKKα/ß, IκBα, and p65) pathways activation, and reducing the expression of key osteoclast differentiation-related regulatory factors (NFATc1, c-Fos, TRAP, c-Src, ß3-Integrin, and cathepsin K) in osteoclast precursors, ultimately negatively regulating osteoclastogenesis. These findings suggest that niloticin could serve as a novel osteoclastogenesis inhibitor and might have beneficial effects on bone health.

Roburic Acid Targets TNF to Inhibit the NF-κB Signaling Pathway and Suppress Human Colorectal Cancer Cell Growth.

Tumor necrosis factor (TNF)-stimulated nuclear factor-kappa B (NF-κB) signaling plays very crucial roles in cancer development and progression, and represents a potential target for drug discovery. Roburic acid is a newly discovered tetracyclic triterpene acid isolated from oak galls and exhibits anti-inflammatory activity. However, whether roburic acid exerts antitumor effects through inhibition of TNF-induced NF-κB signaling remains unknown. Here, we demonstrated that roburic acid bound directly to TNF with high affinity (KD = 7.066 µM), blocked the interaction between TNF and its receptor (TNF-R1), and significantly inhibited TNF-induced NF-κB activation. Roburic acid exhibited antitumor activity in numerous cancer cells and could effectively induce G0/G1 cell cycle arrest and apoptosis in colorectal cancer cells. Importantly, roburic acid inhibited the TNF-induced phosphorylation of IKKα/ß, IκBα, and p65, degradation of IκBα, nuclear translocation of p65, and NF-κB-target gene expression, including that of XIAP, Mcl-1, and Survivin, in colorectal cancer cells. Moreover, roburic acid suppressed tumor growth by blocking NF-κB signaling in a xenograft nude mouse model of colorectal cancer. Taken together, our findings showed that roburic acid directly binds to TNF with high affinity, thereby disrupting its interaction with TNF-R1 and leading to the inhibition of the NF-κB signaling pathway, both in vitro and in vivo. The results indicated that roburic acid is a novel TNF-targeting therapeutics agent in colorectal cancer as well as other cancer types.

(-)-Epigallocatechin-3-gallate inhibits osteoclastogenesis by blocking RANKL-RANK interaction and suppressing NF-κB and MAPK signaling pathways.

Consuming green tea has many health benefits, including regulating bone metabolism and ameliorating osteoporosis, mainly in older and postmenopausal women. This osteoprotective effect has been attributed to the biologically active polyphenol (-)-epigallocatechin-3-gallate (EGCG). Although EGCG inhibits osteoclastogenesis, its underlying molecular mechanism remains to be elucidated. Interaction between receptor activator of nuclear factor (NF)-κB ligand (RANKL) and RANK plays critical roles in the differentiation and activation of osteoclasts and is therefore considered a therapeutic target for osteoclast-related diseases such as osteoporosis. In the present study, we found that EGCG can bind directly to RANK and RANKL and interfere with their interaction, thereby suppressing RANKL-induced phosphorylation of IKKα/ß, IκBα, p65, JNK, ERK1/2, and p38 and key downstream regulatory factors, including nuclear factor of activated T cell c1 (NFATc1), c-Fos, tartrate-resistant acid phosphatase (TRAP), c-Src, and cathepsin K, in osteoclast precursors. This can ultimately inhibit osteoclastogenesis. Taken together, our results show that EGCG can bind directly to RANK and RANKL and block their interaction and that, by inhibiting NF-κB and mitogen-activated protein kinase (MAPK) signaling pathways, it negatively regulates RANKL-induced osteoclastogenesis in RAW 264.7 cells. Thus, regular consumption of EGCG in green tea can inhibit the development and progression of osteoclast-related diseases.

Acanthopanax senticosus aqueous extract ameliorates ovariectomy-induced bone loss in middle-aged mice by inhibiting the receptor activator of nuclear factor-κB ligand-induced osteoclastogenesis.

Acanthopanax senticosus (Ciwujia) has broad-spectrum pharmacological activities, including osteoprotective effects. However, the mechanisms underlying these effects remain unclear. We investigated whether Acanthopanax senticosus aqueous extract (ASAE) ameliorates ovariectomy-induced bone loss in middle-aged mice through inhibition of osteoclastogenesis. In vitro, ASAE significantly suppressed the receptor activator of nuclear factor-κB ligand (RANKL)-stimulated osteoclast differentiation and formation of F-actin rings by downregulating the expression of the nuclear factor of activated T cells, cytoplasmic 1 (NFATc1), c-Fos, and osteoclastogenesis-related marker genes and proteins, including c-Src, tartrate-resistant acid phosphatase (TRAP), cathepsin K, ß3-integrin, and matrix metallopeptidase-9 (MMP-9). This was achieved by inhibiting RANK signaling pathways, including p65, c-Jun N-terminal kinase, extracellular signal-regulated kinase, and p38 in osteoclast precursors. In vivo, ASAE markedly ameliorated bone loss in ovariectomized (OVX) middle-aged mice. ASAE significantly inhibited the serum levels of tartrate-resistant acid phosphatase 5b (TRACP-5b) and RANKL, whereas it increased those of osteocalcin, procollagen 1 N-terminal peptide (P1NP), and osteoprotegerin in OVX mice. ASAE significantly inhibited the OVX-induced expression of osteoclast-specific proteins and genes in the femur. In conclusion, ASAE prevents ovariectomy-induced bone loss in middle-aged mice by inhibiting RANKL-induced osteoclastogenesis through suppression of RANK signaling pathways and could be potentially used in mediated treatment of osteoclast-related diseases (e.g., osteoporosis).

Caffeine Targets G6PDH to Disrupt Redox Homeostasis and Inhibit Renal Cell Carcinoma Proliferation.

Glucose-6-phosphate dehydrogenase (G6PDH) is the rate-limiting enzyme in the pentose phosphate pathway (PPP) and plays a crucial role in the maintenance of redox homeostasis by producing nicotinamide adenine dinucleotide phosphate (NADPH), the major intracellular reductant. G6PDH has been shown to be a biomarker and potential therapeutic target for renal cell carcinoma (RCC). Here, we report a previously unknown biochemical mechanism through which caffeine, a well-known natural small molecule, regulates G6PDH activity to disrupt cellular redox homeostasis and suppress RCC development and progression. We found that caffeine can inhibit G6PDH enzymatic activity. Mechanistically, caffeine directly binds to G6PDH with high affinity (K D = 0.1923 µM) and competes with the coenzyme NADP+ for G6PDH binding, as demonstrated by the decreased binding affinities of G6PDH for its coenzyme and substrate. Molecular docking studies revealed that caffeine binds to G6PDH at the structural NADP+ binding site, and chemical cross-linking analysis demonstrated that caffeine inhibits the formation of dimeric G6PDH. G6PDH inhibition abrogated the inhibitory effects of caffeine on RCC cell growth. Moreover, inhibition of G6PDH activity by caffeine led to a reduction in the intracellular levels of NADPH and reactive oxygen species (ROS), and altered the expression of redox-related proteins in RCC cells. Accordingly, caffeine could inhibit tumor growth through inhibition of G6PDH activity in vivo. Taken together, these results demonstrated that caffeine can target G6PDH to disrupt redox homeostasis and inhibit RCC tumor growth, and has potential as a therapeutic agent for the treatment of RCC.

Ellagic acid blocks RANKL-RANK interaction and suppresses RANKL-induced osteoclastogenesis by inhibiting RANK signaling pathways.

Ellagic acid (EA) is a naturally occurring polyphenolic compound that has been shown to exhibit diverse beneficial pharmacological activities including anti-osteoclastogenesis effect. However, the molecular mechanism by which EA inhibits osteoclastogenesis remains to be elucidated. The protein-protein interaction between receptor activator of nuclear factor (NF)-κB ligand (RANKL) and its receptor RANK contributes to osteoclast differentiation and activation in bone remodeling, and is regarded as an important therapeutic target for the treatment of osteoporosis. The current study is focused on investigating whether EA can directly bind to RANKL and/or RANK and block the interaction between RANKL and RANK, thereby inhibiting downstream signaling pathways. Interestingly, we found that EA had strong affinities to RANK and RANKL, with the estimated equilibrium dissociation constants (KD) of 2.485 × 10-11 and 1.688 × 10-9 M, respectively, and could disrupt the interaction between RANKL and RANK, thereby inhibiting RANKL-induced canonical RANK signaling pathways (p65, JNK, ERK, and p38) and expression of downstream master transcriptional factors (NFATc1 and c-Fos) and osteoclast-specific genes and proteins (TRAP, c-Src, and cathepsin K), which could ultimately suppress RANKL-induced osteoclast differentiation and F-actin ring formation. Taken together, our results revealed that EA could block RANKL-RANK interaction and suppress RANKL-induced osteoclastogenesis by inhibiting RANK signaling pathways in RAW 264.7 murine macrophages.

Oxidation derivative of (-)-epigallocatechin-3-gallate (EGCG) inhibits RANKL-induced osteoclastogenesis by suppressing RANK signaling pathways in RAW 264.7 cells.

Tea consumption has positive effects on the skeletal system and prevents postmenopausal osteoporosis, mainly by inhibiting osteoclastogenesis. In green tea, (-)-epigallocatechin-3-gallate (EGCG) is the most abundant and active compound and has been shown to inhibit RANKL-induced osteoclast formation. Taking into account the highly oxidizable and unstable nature of EGCG, we hypothesized that EGCG oxidation product exhibits greater anti-osteoclastogenesis potential than EGCG. In this study, we successfully isolated and identified an EGCG oxidation derivative, (-)-gallocatechin gallate (compound 2), using a chemical oxidation strategy. We then compared the ability of compound 2 and EGCG to inhibit RANKL-induced osteoclastogenesis in RAW 264.7 cells. The results of TRAP staining and F-actin ring immunofluorescent staining showed that compound 2 exhibits stronger inhibition of RANKL-induced osteoclast differentiation and F-actin ring formation, respectively, than EGCG. Additionally, quantitative real-time PCR (qRT-PCR) and western blotting analyses showed that compound 2 significantly and more strongly inhibited the expression of osteoclastogenesis-related marker genes and proteins, including c-Src, TRAP, cathepsin K, ß3-Integrin, and MMP-9, compared with EGCG. Furthermore, compound 2 significantly suppressed RANKL-induced expression of NFATc1 and c-Fos, the master transcriptional regulators of osteoclastogenesis, more strongly than EGCG. Mechanistically, molecular interaction assays showed that compound 2 binds to RANK with high affinity (KD = 189 nM) and blocks RANKL-RANK interactions, thereby suppressing RANKL-induced early RANK signaling pathways including p65, JNK, ERK, and p38 in osteoclast precursors. Taken together, this study demonstrates for the first time that an oxidation derivative of EGCG (compound 2) inhibits RANKL-induced osteoclastogenesis by suppressing RANK signaling pathways in RAW 264.7 cells.

Effect of caffeine on ovariectomy-induced osteoporosis in rats.

Caffeine (1,3,7-trimethylxanthine) is a naturally occurring plant xanthine alkaloid present in many commonly consumed beverages worldwide, including tea, coffee, and cocoa. Although moderate caffeine intake is generally considered to exert positive effects on human health, its effect on bone metabolism remains controversial. The aim of this study was to systematically evaluate the pharmacological effect of long-term administration of caffeine on ovariectomy-induced postmenopausal osteoporosis in female rats. A sham operation or ovariectomy was performed to establish the ovariectomy rat model. The ovariectomized (OVX) rats were divided into five subgroups: OVX with vehicle (model), OVX with raloxifene hydrochloride (RLX, positive control; 4 mg/kg body weight/day), and OVX with low-, medium-, and high-dose caffeine (9.6, 19.2, and 38.4 mg/kg of body weight/day, respectively). Their corresponding treatments were administered intragastrically for 13 weeks. In-vivo studies showed that treatment with caffeine effectively improved the lipid profiles and increased the concentration of calcium in the serum of OVX rats. Medium- or high-dose treatment with caffeine significantly decreased the activities of alkaline and acid phosphatases in OVX rats. In addition, treatment with caffeine (at any dose) did not adversely affect organ weights, organ coefficients, femoral length, bone mineral density, biomechanical properties, or bone microarchitecture in OVX rats. Collectively, our study demonstrated that caffeine did not exert a damaging effect on the skeletal system of OVX rats.

Interactions between ß-cyclodextrin and tea catechins, and potential anti-osteoclastogenesis activity of the (-)-epigallocatechin-3-gallate-ß-cyclodextrin complex.

Galloylated catechins, the most important secondary metabolites in green tea including (-)-epigallocatechin-3-gallate (EGCG) and (-)-epicatechin-3-gallate, constitute nearly 75% of all tea catechins and have stronger health effects than non-galloylated catechins such as (-)-epigallocatechin and (-)-epicatechin. EGCG is the most abundant, active, and thoroughly investigated compound in green tea, and its bioactivity might be improved by complexing with ß-cyclodextrin (ß-CD). We investigated interactions between four catechins and ß-CD in a PBS buffer solution of pH 6.5 at 25 °C using biolayer interferometry and isothermal titration calorimetry, and to determine whether ß-CD could enhance the anti-osteoclastogenesis effect of EGCG. ß-CD could directly bind galloylated catechins at a stoichiometric ratio close to 1 : 1, with high specificities and affinities, and these inclusion interactions were primarily enthalpy-driven processes. We synthesized the EGCG-ß-CD complex and identified it using infrared radiation and nuclear magnetic resonance spectra. Interestingly, we revealed that the EGCG-ß-CD complex could inhibit osteoclastogenesis significantly more than EGCG.

Theabrownin suppresses in vitro osteoclastogenesis and prevents bone loss in ovariectomized rats.

Drinking tea exhibits beneficial effects on bone health and may protect against osteoporosis, particularly in postmenopausal women. Theabrownin (TB) is the main component responsible for the biological activities of Pu-erh tea, but whether it possesses anti-osteoporotic potential remains unknown. Here we investigated the in vitro and in vivo anti-osteoporotic effects of TB in the RAW 264.7 cell line and ovariectomized (OVX) rats, respectively. Our in vitro studies showed that TB significantly suppressed RANKL-induced osteoclastogenesis and the expression of related marker proteins, including NFATc1, TRAP, c-Fos, and cathepsin K. In vivo studies showed that TB treatment effectively ameliorated blood biochemical parameters, organ weights and organ coefficients in OVX rats. In addition, TB treatment significantly improved femoral bone mineral density (BMD) and biomechanical properties. What's more, TB treatment strikingly ameliorated bone microarchitecture in OVX rats because of increased cortical bone thickness and trabecular bone area in the femur. Our study therefore demonstrated that TB can inhibit RANKL-induced osteoclastogenesis in vitro and prevent bone loss in ovariectomized rats. Consequently, TB has a promising potential in postmenopausal osteoporosis treatment.

Tea polysaccharide inhibits RANKL-induced osteoclastogenesis in RAW264.7 cells and ameliorates ovariectomy-induced osteoporosis in rats.

BACKGROUND AND PURPOSE: Tea drinking has positive effects on bone health and may prevent and treat osteoporosis, especially in older and postmenopausal women. Tea polysaccharide (TPS) is a major bioactive constituent in tea. Despite its profound effects on human health, whether TPS has anti-osteoporotic effects remains largely unknown. As such, we investigated the anti-osteoporotic effects of TPS. METHODS: In vitro, TPS effects on osteoclastogenesis were examined using osteoclast precursor RAW264.7 cells. TPS effects on osteoclastogenesis-related expression of marker genes and proteins were determined by gene expression and immunoblotting analyses, respectively. For in vivo studies, 12-week-old female Wistar rats were divided randomly into a sham-operated group (sham) and four ovariectomized (OVX) subgroups: OVX with vehicle (model) and OVX with low-, medium-, and high-dose TPS (0.32, 0.64 and 1.28 g/kg body weight/day, respectively). TPS was administered intragastrically to rats for 13 weeks. Body weight, blood biochemical parameters, organ weight, organ coefficients, femoral length, bone mineral density (BMD), biomechanical properties, and bone microarchitecture were documented. RESULTS: TPS inhibited osteoclast differentiation significantly and dose-dependently, and its inhibitory effect was not due to toxicity to RAW264.7 cells. TPS suppressed expression of osteoclastogenesis-related marker genes and proteins significantly. In in vivo studies, medium-dose TPS treatment ameliorated OVX-induced calcium loss significantly. Low-dose TPS treatment decreased the activity of acid phosphatase (ACP) in OVX rats significantly. In addition, TPS treatment improved other blood biochemical parameters and femoral biomechanical properties to a certain extent. More importantly, TPS treatment ameliorated bone microarchitecture in OVX rats strikingly because of increased cortical bone thickness and trabecular bone area in the femur. CONCLUSION: TPS can inhibit receptor activator nuclear factor-kappa B ligand (RANKL)-induced osteoclastogenesis in RAW264.7 cells and ameliorate ovariectomy-induced osteoporosis in rats.

Pu-erh Tea Extract Ameliorates Ovariectomy-Induced Osteoporosis in Rats and Suppresses Osteoclastogenesis In Vitro.

Background and Objective: Tea drinking is associated with positive effects on bone health and may protect against osteoporosis, especially in elderly women. Pu-erh tea has many beneficial effects on human health; however, whether Pu-erh tea has anti-osteoporotic potential remains unclear. Thus, we investigated the effects of Pu-erh tea extract (PTE) on ovariectomy-induced osteoporosis in rats and on osteoclastogenesis in vitro. Methods: Female Wistar rats were divided into six groups: the sham, model, and Xian-Ling-Gu-Bao capsule (XLGB) groups, and the low-, medium-, and high-dose PTE groups. Ovariectomized (OVX) rats were used as an animal model of osteoporosis. The animals were intragastrically administered distilled water, XLGB, or different concentrations of PTE for 13 weeks. Body weight, blood biochemical indicators, relative organ coefficients, femoral bone mineral density (BMD), bone biomechanical properties, and bone microarchitecture were examined and analyzed. Additionally, the in vitro effects of PTE on osteoclastic activities were investigated using the RAW 264.7 cell line as an osteoclast differentiation model. The effects of PTE on osteoclast differentiation and the expression of osteoclast-specific genes and proteins were determined. Results: PTE reduced OVX-induced body weight gain after 6 weeks of treatment, and the high-dose exerted a significant effect. High-dose PTE significantly ameliorated OVX-induced estradiol (E2) deficiency. PTE treatment maintained calcium and phosphorus homeostasis and improved other blood biochemical parameters to various degrees. In addition, PTE treatment improved organ coefficients of the femur, uterus, and vagina and improved femoral BMD and bone biomechanical properties. PTE treatment strikingly ameliorated bone microarchitecture. Moreover, in the in vitro studies, osteoclast differentiation using the differentiation cell model was significantly inhibited by PTE without cytotoxic effects. Additionally, PTE efficaciously suppressed the expression of key osteoclast-specific genes and proteins. Conclusion: PTE can ameliorate ovariectomy-induced osteoporosis in rats and suppress osteoclastogenesis in vitro.

Structural characterization and immunomodulating activity of polysaccharide from Dendrobium officinale.

A neutral heteropolysaccharide (DOP-1-1) consisted by mannose and glucose (5.9:1) with an average molecular weight at about 1.78×10(5) Da was purified from Dendrobium officinale. Based on Fourier transform infrared spectrum (FT-IR) and nuclear magnetic resonance (NMR) spectra, it suggested that partial structure of DOP-1-1 is an O-acetylated glucomannan with ß-d configuration in pyranose sugar forms. The immunomodulatory activity of DOP-1-1 was evaluated by secretion level of cytokine (interleukin (IL)-1ß and IL-10) and tumor necrosis factor (TNF)-α in vitro. Our results suggested that DOP-1-1 could stimulate cytokine production (TNF-α, IL-1ß) in cells. These findings demonstrated that the purified polysaccharide from D. officinale presented significant immune-modulating activities. Furthermore, by Western-blot we can found that the signaling pathways of DOP-1-1 induced immune activities involving ERK1/2 and NF-кB. As to antioxidant activity, DOP-1-1 hadn't showed remarkable scavenging capacity of 1,1-diphenyl-2-picrylhydrazyl radical (DPPH) in contrast with other studies of polysaccharides from D. officinale.