To study the anti-inflammatory activity of glycyrrhizin acid, ligustrazine and puerarin. In the study, the liquichip-based high-throughput synchronous detection technique for 23 inflammatory factors, uniform design, comprehensive weight method were adopted to study the effect of different combined administration of glycyrrhizin acid, ligustrazine and puerarin in inhibiting the expression of lipopolysaccharide (LPS)-induced RAW264. 7 cells and multiple inflammatory cytokines. In the study, the uniform design table U₉ (9³) was adopted to design doses of glycyrrhizin acid, ligustrazine and puerarin. The liquichip technique was used to detect the effect of different combined administration of glycyrrhizin acid, ligustrazine and puerarin on the 23 cytokines expressed in LPS-induced mouse macrophage RAW264. 7 inflammation model. The traditional Chinese medicine component optimization software and the improved least angle regression algorithm were used to analyze the dose-effect relationship among the three components and the cytokine inhibition rate and produce the regression equation. The comprehensive weight method was applied to get the optimal dose ratio of glycyrrhizic acid, ligustrazine and puerarin with highest efficacy of 25:2:13 and verify the optimal dose ratio. The verification results were consistent with the prediction trend, indicating the accuracy of the mathematical model for predicting the experiment. The experimental results showed the multi-target and multi-level efficacies of glycyrrhizic acid, ligustrazine and puerarin and the high anti-inflammatory activity of their combined administration, which provides powerful basis for subsequent drug development.
Animals ; Anti-Inflammatory Agents ; pharmacology ; Cytokines ; Glycyrrhizic Acid ; pharmacology ; Isoflavones ; pharmacology ; Lipopolysaccharides ; immunology ; Macrophages ; drug effects ; immunology ; Mice ; NF-kappa B ; genetics ; immunology ; Pyrazines ; pharmacology ; RAW 264.7 Cells

OBJECTIVETo investigate the effect of Thymosin and growth hormone(GH) on inflammatory response in burn rats or burn rats with sepsis.

METHODSSixty-four SD rats were randomly divided into normal control group (NC, without treatment), sepsis group (S, with injection of LPS), sepsis + Thymosin group (ST, with successive injection of Thymosin and LPS), sepsis + GH group [SGH, with successive injection of recombinant human GH (rhGH) and LPS], burn group, burn + sepsis group (BS, with injection of LPS after burn), burn + sepsis + Thymosin group (BST, with successive injection of Thymosin and LPS after burn), burn + sepsis + GH (BSGH, with successive injection of rhGH and LPS after burn), with 8 rats in each group. Specimens of spleen tissues were harvested to determine HLA-DR in lymphocyte and evaluate inflammatory cell infiltration (score). Specimens of peripheral blood were collected to determine Toll-like receptor 4 (TLR4) level in monocyte and serum level of TNF-alpha, IL-4, IL-6, IL-10.

RESULTSCompared with those in NC group, serum level of IL-10 in S group decreased obviously, while other indices increased obviously (P < 0.01). The levels of HLA-DR and TLR4 and serum level of TNF-alpha were similar between SGH and ST groups (P > 0.05). Compared with those in SGH group [(2.87 +/- 0.04) score, and IL-6 (0.0083 +/- 0.0018) microg/mg, IL-4 (0.0102 +/- 0.0021) microg/mg, IL-10 (0.0310 +/- 0.0027) microg/mg, respectively], degree of inflammatory cell infiltration (1.50 +/- 0.76) score and serum levels of IL-6, IL-4, IL-10 of rats in ST group decreased obviously (0.0064 +/- 0.0012, 0.0058 +/- 0.0024, 0.0230 +/- 0.0021 microg/mg, respectively, P < 0.01). The levels of HLA-DR, TLR4 and inflammatory cell infiltration degree of spleen in B group were respectively higher than those in NC group and lower than those in BS group. Compared with those in NC group, serum levels of TNF-alpha, IL-6 in B group increased significantly, while IL-4, IL-10 showed an opposite tendency. There was no obvious difference between BST and BSGH groups in serum levels of HLA-DR and IL-6 (P > 0.05). Compared with those in BST group, inflammatory cell infiltration degree in spleen and the levels of TLR, TNF-alpha obviously decreased (P < 0.01), while IL-4 and IL-10 levels increased in BSGH group (P < 0.01).

CONCLUSIONSInhibitive effects between Thymosin and GH on extensive inflammatory reaction were similar with or without trauma, and GH has better effect as compared with Thymosin when with trauma.

Animals ; Anti-Inflammatory Agents ; pharmacology ; Burns ; immunology ; Human Growth Hormone ; pharmacology ; Inflammation ; immunology ; Male ; Rats ; Rats, Sprague-Dawley ; Sepsis ; immunology ; Thymosin ; pharmacology

The saponin ginsenoside Rk1 is a major compound isolated from ginseng. Ginsenoside Rk1 has been reported to have anti-inflammatory and anti-tumor properties and to be involved in the regulation of metabolism. However, the effect and mechanism of anti-inflammatory action of ginsenoside Rk1 has not been fully clarified. We investigated whether ginsenoside Rk1 could suppress the inflammatory response in lipopolysaccharide-stimulated RAW264.7 macrophages and to explore its mechanism of the action. RAW264.7 cells were treated with LPS (1 μg·mL) in the absence or the presence of Ginsenoside Rk1 (10, 20, and 40 μmol·L). Then the inflammatory factors were tested with Griess reagents, ELISA, and RT-PCR. The proteins were analyzed by Western blotting. Ginsenoside Rk1 inhibited lipopolysaccharide-induced expression of nitric oxide (NO), interleukin (IL)-6, IL-1β, tumor necrosis factor (TNF)-α, and monocyte chemotactic protein (MCP)-1. Ginsenoside Rk1 inhibited the lipopolysaccharide-stimulated phosphorylation of NF-κB and janus kinase (Jak)2 and signal transducer and activator of transcription (Stat)3 at Ser727 and Tyr705. These data suggested that ginsenoside Rk1 could inhibit expression of inflammatory mediators and suppress inflammation further by blocking activation of NF-κB and the Jak2/Stat3 pathway in LPS-stimulated RAW264.7 cells.
Animals ; Anti-Inflammatory Agents ; pharmacology ; Ginsenosides ; pharmacology ; Interleukin-6 ; genetics ; immunology ; Janus Kinase 2 ; genetics ; immunology ; Lipopolysaccharides ; pharmacology ; Macrophages ; drug effects ; immunology ; Mice ; RAW 264.7 Cells ; STAT3 Transcription Factor ; genetics ; immunology ; Tumor Necrosis Factor-alpha ; genetics ; immunology

OBJECTIVE: To investigate anti-inflammatory and immunomodulation effects of different ecotype from Isatidis Radix growing in Gansu province. METHODS: Mice were randomly divided into 6 groups (=11)and used the auricular swelling and paw edema to observe the anti-inflammatory effects of Isatidis Radix; Mice were randomly divided into 7 groups (=11) and through the gasbag synovitis model to observe the anti-inflammatory effects of Isatidis Radix; Mice were randomly divided into 6 groups (=11), the immunosuppressed model were established by injection of cyclophosphamide (CTX) to study the effects of Isatidis Radix on index of thymus, blood routine and cytokines. RESULTS: Gansu different ecotype from Isatidis Radix could reduce the swelling of the mice auricle, paw edema and total protein, leukotriene B(LTB)and malonaldehyde(MDA) in airbag synovitis exudates, and upgrade serum levels of superoxide dismutase (SOD); Degrade the tumor necrosis factor-α (TNF-α) and upgrade the index of thymus, the number of red and white corpuscles, the level of interferon-γ (IFN-γ), interleukin-4 (IL-4) (<0.05, 0.01) of mice immunosuppressed model; Above the research of anti-inflammatory and immunomodulation, there were no significant differences between Isatidis Radix of Gansu different ecotype and tetraploid. CONCLUSIONS Different ecotype of Isatidis Radix has obvious functions in anti-inflammatory and immunomodulation, but there are no significant differences between Gansu different ecotype and tetraploid.
Animals ; Anti-Inflammatory Agents ; pharmacology ; China ; Cytokines ; immunology ; Drugs, Chinese Herbal ; pharmacology ; Ecotype ; Immunomodulation ; drug effects ; Isatis ; chemistry ; Mice ; Plant Extracts ; pharmacology ; Random Allocation

OBJECTIVETo study and compare the anti-inflammatory effect and molecular mechanism of artemisinin and dihydroartemisinin.

METHODMouse mononuclear macrophage RAW264.7 cells were stimulated to release inflammatory mediators such as TNF-alpha, IL-6 and NO, in order to assess the drugs' inhibitory effect on macrophage's release of above inflammatory mediators. The levels of TNF-alpha and IL-6 were determined by ELISA and the cytotoxicity was determined by MTT method. The protein expression of iNOS, COX-2 and beta-actin were tested by Western blot. The enzymatic activity of COX-2 was determined by colorimetric method.

RESULTDihydroartemisinin significantly inhibited LPS-induced release of TNF-alpha, IL-6 and NO from RAW264.7 in mice with the concentration range of 12.5 - 100 micromol x L(-1), and showed good dose dependence. Artemisinin only inhibited the IL-6 release to a certain extent.

CONCLUSIONDihydroartemisinin inhibits macrophages from releasing inflammatory factors TNF-alpha and IL-6 and inflammatory mediators NO by down-regulating iNOS protein. Artemisinin may help dihydroartemisinin to show its anti-inflammatory effect through metabolism.

Animals ; Anti-Inflammatory Agents ; pharmacology ; Artemisinins ; pharmacology ; Cell Line ; Gene Expression ; drug effects ; Inflammation Mediators ; immunology ; Interleukin-6 ; genetics ; immunology ; Macrophages ; drug effects ; immunology ; Mice ; Nitric Oxide ; immunology ; Tumor Necrosis Factor-alpha ; genetics ; immunology

OBJECTIVETo examine the anti-inflammatory mechanism of total flavonoids of Glycyrrhizae Radix et Rhizoma (TFGR) and its ingredient on IFN-gamma and LPS-induced macrophage RAW264.7.

METHODSolvent extraction and macroporous resin enrichment were adopted for preparing ethanol extracts of Glycyrrhizae Radix et Rhizoma, components and total flavonoids. Ultraviolet spectroscopy was used to determine the content of total flavonoids. An IFN-gamma and LPS-induced cell inflammatory model was established. Griess reaction was used for detecting the effect of extracts at all levels and flavonoid monomers on nitrite content in cell culture supernatant. FRAP was used for measuring anti-oxidation capacity. RT-PCR was used for determining the effect of TFGR and isoliquiritigenins on intracellular inducible nitric oxide synthase iNOS, COX-2, IL-6 and PPAR-gamma. Western blot was used for detecting the effect of TFGR and isoliquiritigenins on iNOS, COX-2 and MAPK signal transduction pathways.

RESULTCompared with other extracts, ethyl acetate fractions from Glycyrrhizae Radix et Rhizoma showed the highest inhibition ratio on nitrite content at the same concentration. After being enriched with macroporous resin, TFGR (60. 08% of liquiritin) of ethyl acetate extracts from Glycyrrhizae Radix et Rhizoma showed dose-dependence, and inhibited the nitrite content in cell culture supernatant, which was superior to ethyl acetate extracts, and had the protective effect on post-stimulated cell activity, with a stronger total anti-oxidation than other extracts. TFGR inhibited iNOS, IL-6 mRNA, protein expressions of iNOS, COX-2 and IL-6. Isoliquiritigenin, a flavonoid monomer, could inhibited iNOS, COX-2 gene and protein expression and gene expressions of IL-1beta and IL-6, and upside-regulated gene expression of PPAR-gamma.

CONCLUSIONActivity-oriented extraction suggests that ethyl acetate fractions from Glycyrrhizae Radix et Rhizoma is one of components with anti-inflammatory activity. TFGR obtained by enriching the active component showed dose-dependence, and inhibited the nitrite content in cell culture supernatant. The anti-inflammatory effect is partially achieved by regulating ERK signal pathway and inhibiting iNOS and COX-2 gene and protein expressions through extracellular signals of mitogen activated protein kinases (MAPKs). Specifically, isoliquiritigenin may be a component with TFGR anti-inflammatory activity.

Animals ; Anti-Inflammatory Agents ; pharmacology ; Cell Line ; Drugs, Chinese Herbal ; pharmacology ; Flavonoids ; pharmacology ; Glycyrrhiza ; chemistry ; Interleukin-1beta ; genetics ; immunology ; Interleukin-6 ; genetics ; immunology ; Macrophages ; drug effects ; immunology ; Mice ; NF-kappa B ; genetics ; immunology ; Rhizome ; chemistry

Multi-components in herbal formulae exert holistic effects in synergistic or additive manners. However, appropriate strategies and supportive evidences are still lacking to uncover the synergistic or additive combinations. The present investigation aimed at seeking a screening strategy to identify the targeted combinations in GuGe FengTong Tablet (GGFTT), an herbal formula. Two compounds, belonging to different chemical classes, were combined with different concentration ratios and their anti-inflammation effects were investigated. The most significant anti-inflammatory combinations were evaluated by combination index (CI) method (additive effect, CI = 1; synergism, CI < 1; antagonism, CI > 1). The modulating effects of candidate combinations on pro-inflammatory cytokines and MAPKs signaling pathway were also detected. Two combinations, "biochanin A + 6-gingerol" (Bio-6G) and "genistein + 6-gingerol" (Gen-6G), showed synergistic effects (CI < 1), and Bio-6G was selected for further study. Compared with single compound, Bio-6G could synergistically inhibit the production of pro-inflammatory cytokines (TNF-α, IL-1β, and IL-6) and the activation of MAPKs signaling pathway in LPS-stimulated RAW264.7 cells. The combined results showed that Bio-6G was a synergistic anti-inflammatory combination in GGFTT. Our results could provide a useful strategy to screen the synergistic combinations in herbal formulae.
Animals ; Anti-Inflammatory Agents ; chemistry ; pharmacology ; Drug Compounding ; Drug Synergism ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; Interleukin-1beta ; immunology ; Macrophages ; drug effects ; immunology ; Mice ; NF-kappa B ; immunology ; RAW 264.7 Cells ; Tablets ; chemistry ; pharmacology ; Tumor Necrosis Factor-alpha ; immunology

The purpose of this study was to identify the effect of sildenafil citrate on IL-1 beta induced nitric oxide (NO) synthesis and iNOS expression in human synovial sarcoma SW982 cells. IL-1 beta stimulated the cells to generate NO in both dose- and time-dependent manners. The IL-1 beta -induced NO synthesis was inhibited by guanylate cyclase (GC) inhibitor, LY83583. When the cells were treated with 8-bromo-cGMP, a hydrolyzable analog of cGMP, NO synthesis was increased upto 5-fold without IL-1 beta treatment suggesting that cGMP is an essential component for increasing the NO synthesis. Synoviocytes and chondrocytes contain strong cGMP phosphodiesterase (PDE) activity, which has biochemical features of PDE5. When SW982 cells were pretreated with sildenafil citrate (Viagra), a PDE5 specific inhibitor, sildenafil citrate significantly inhibited IL-1 beta -induced NO synthesis and iNOS expressions. From this result, we noticed that PDE5 activity is required for IL-1 beta -induced NO synthesis and iNOS expressions in human synovial sarcoma cells, and sildenafil citrate may be able to suppress an inflammatory reaction of synovium through inhibition of NO synthesis and iNOS expression by cytokines.
Anti-Inflammatory Agents/immunology/pharmacology ; Cell Line, Tumor ; Cyclic GMP/analogs & derivatives/immunology/metabolism ; Cyclic Nucleotide Phosphodiesterases, Type 2/antagonists & inhibitors/metabolism ; Humans ; Interleukin-1beta/*metabolism ; Male ; Nitric Oxide/*biosynthesis/genetics/immunology ; Nitric Oxide Synthase Type II/*biosynthesis/genetics/immunology ; Phosphodiesterase Inhibitors/immunology/*pharmacology ; Piperazines/immunology/*pharmacology ; Purines/immunology/pharmacology ; Signal Transduction/drug effects/genetics/immunology ; Sulfones/immunology/*pharmacology ; Synovial Membrane/enzymology/immunology

This study aims to investigate the PPARγ agonists isolated from the aqueous extract of Siegesbeckia pubescens( SPA) and their anti-inflammatory activities in vitro. The 293 T cells transfected transiently with PPARγ recombinant plasmid were used as a screening model to guide the isolation of PPARγ activitating components,and then PPARγ activities were measured by double luciferase reporter gene assay. The chemical structures were identified by chromatography or spectroscopic techniques. Furthermore,a UC inflammatory model in vitro was established on HT-29 cells by stimulating with TNF-α． The mRNA levels and secretion of proinflammatory cytokines on HT-29 cells,such as IL-1β,TNF-α,IL-8,were detected by RT-PCR and ELISA. The results showed that five diterpenoids were obtained from the fraction D_(50) with the strongest PPARγ activity among others in SPA,and determined as kirenol( 1),darutigenol( 2),enantiomeric-2-ketone-15,16,19-three hydroxypinomane-8( 14)-ene-19-O-β-D-glucoside( 3),darutoside( 4),enantiomeric-2-β,15,16,19-four hydroxypinomane-8( 14)-ene-19-O-β-D-glucoside( 5),respectively. All the compounds exhibited active effects on PPARγ in a concentration-dependent manner( P<0. 01). In addition,compound 1 significantly inhibited the expression of IL-1β mRNA and secretion of IL-8 on HT-29 cells inflammation model( P<0. 001); both compounds 2 and 3 effectively inhibited the expression of IL-1β,TNF-α,IL-8 mRNA and secretion of IL-8( P<0. 01 or P<0. 001),although at different extent; compound 4 significantly inhibited the expression of IL-1β and TNF-α mRNA( P<0. 01 or P<0. 001),while compound 5 inhibited the expression of IL-1β mRNA obviously( P<0. 001). In conclusion,the diterpenoids 1-5 isolated from S. pubescens have the PPARγ activation activities and potential effects of anti-UC in vitro.
Anti-Inflammatory Agents/pharmacology* ; Asteraceae/chemistry* ; Colitis, Ulcerative ; Cytokines/immunology* ; Diterpenes/pharmacology* ; HT29 Cells ; Humans ; PPAR gamma/agonists* ; Tumor Necrosis Factor-alpha

The aim of this study was to investigate the anti-inflammatory effect of Guizhi Fuling capsule and its active complex (consistent of 15 active compounds) on LPS-induced RAW264. 7 cells. The effect of Guizhi Fuling capsule and its active complex on cell viability in RAW264. 7 cells were determined by MTT assay. The inhibitory effect of Guizhi Fuling capsule and active complex on the releasing of IL-1β, TNF-α and PGE2 induced by LPS in RAW264. 7 cells was detected by ELISA assay. The expression of IL-1β and mPGES-1 in Guizhi Fuling capsule or active complex treated RAW264. 7 cells was examined by Western blot assay. Guizhi Fuling capsule and active complex showed no significant effect on the cell viability in RAW264. 7 cells at doses range from 12.5 to 400 mg x L(-1). Compared with LPS treated group, Guizhi Fuling capsule and active complex dose dependently reduced the releasing of IL-1β, TNF-α and PGE2 induced by LPS in RAW264. 7 cells. Moreover, the expression of IL-1β and mPGES-1 was decreased after Guizhi Fuling capsule and active complex treatment, which might contribute to the inhibitory effect of Guizhi Fuling capsule in the releasing of IL-1β, TNF-α and PGE2. This study provided the evidence that Guizhi Fuling capsule and active complex remarkably inhibited the releasing of IL-1β, TNF-α and PGE2induced by LPS in RAW264. 7 cells by reducing the expression IL-1β and mPGES-1. This study provided an experimental basis of Guizhi Fuling capsule for the treatment of inflammation and a theoretical basis for the development of effective compounds of Guizhi Fuling capsule.
Animals ; Anti-Inflammatory Agents ; pharmacology ; Cell Line ; Cell Survival ; drug effects ; Drugs, Chinese Herbal ; pharmacology ; Inflammation ; immunology ; Interleukin-1beta ; immunology ; Macrophages ; drug effects ; immunology ; Mice ; Tumor Necrosis Factor-alpha ; immunology