METHODS: Cells were pre-incubated with 32µM of 15dPGJ2 and stimulated with 1ng/mL of IL-1β as an in vitro model of inflammation. Western immunoblotting was used to detect phosphorylated p-65 and phosphorylated c-Jun as markers of NF-κB and AP-1 activation, respectively. mRNA expression of the pro-inflammatory cytokines IL-6, IL-8, and TNF-α was examined, and protein expression of COX-2 and PGE2 were detected by western immunoblotting and ELISA respectively. Myometrial contractility was examined ex-vivo using a myograph.
RESULTS: 15dPGJ2 inhibited IL-1β-induced activation of NF-κB and AP-1, and expression of IL-6, IL-8, TNF-α, COX-2 and PGE2 in myocytes, with no effect on myometrial contractility or cell viability. Despite inhibiting IL-1β-induced activation of NF-κB, expression of IL-6, TNF-α, and COX-2, 15dPGJ2 led to activation of AP-1, increased production of PGE2 and increased cell death in VECs and AECs.
CONCLUSION: We conclude that 15dPGJ2 has differential effects on inflammatory modulation depending on cell type and is therefore unlikely to be a useful therapeutic agent for the prevention of preterm birth.
AIM OF THE STUDY: However, so far there is no literature available on the anti-inflammatory activity of this species. Henceforth, based on the above background and our previous laboratory findings, we hypothesize that phytoconstituents of A. elliptica could possess anti-inflammatory potential against inflammatory mediators including prostaglandin-E2 (PGE2), cyclooxegenase-2 (COX-2) and cytokines (IL-1β and IL-6).
MATERIALS AND METHODS: Vacuum and column chromatography techniques were employed for the isolation of phytoconstituents. The structure elucidation was carried out using HRESI-MS, 1H and 13C-NMR analysis and compared with the published literature. For cytotoxicity analysis, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was performed on peripheral blood mononuclear cells. In-vitro anti-inflammatory activities were evaluated against the levels of PGE2, COX-2, IL-1β and IL-6 in lipopolysaccharide (LPS)-induced human plasma using enzyme-linked immunosorbent assay and radioimmunoassay.
RESULTS: Unprecedentedly, chromatographic purification of methanolic leaves extract afforded five flavones namely vitexin, isovitexin, orientin, isoorientin, schaftoside with three flavanols; kaempferol, myricetin and rutin from A elliptica. In cell viability analysis, isolates did not present cytotoxicity up to 50 μM. In anti-inflammatory evaluation, orientin and isoorientin exhibited strong (≥70%), while isovitexin and vitexin produced strong to moderate (50-69%) PGE2, COX-2, IL-1β and IL-6 inhibition at 25 and 50 μM. Isoorientin, orientin, isovitexin, and vitexin showed significant (p
AIM OF THE STUDY: The molecular mechanisms of the anti-inflammatory properties of M. accedens are not yet understood. Therefore, we examined those mechanisms using a methanol extract of M. accedens (Ma-ME) and determined the target molecule in macrophages.
MATERIALS AND METHODS: We evaluated the anti-inflammatory effects of Ma-ME in lipopolysaccharide (LPS)-stimulated RAW264.7 cells and in an HCl/EtOH-triggered gastritis model in mice. To investigate the anti-inflammatory activity, we performed a nitric oxide (NO) production assay and ELISA assay for prostaglandin E2 (PGE2). RT-PCR, luciferase gene reporter assays, western blotting analyses, and a cellular thermal shift assay (CETSA) were conducted to identify the mechanism and target molecule of Ma-ME. The phytochemical composition of Ma-ME was analyzed by HPLC and LC-MS/MS.
RESULTS: Ma-ME suppressed the production of NO and PGE2 and the mRNA expression of proinflammatory genes (iNOS, IL-1β, and COX-2) in LPS-stimulated RAW264.7 cells without cytotoxicity. Ma-ME inhibited NF-κB activation by suppressing signaling molecules such as IκBα, Akt, Src, and Syk. Moreover, the CETSA assay revealed that Ma-ME binds to Syk, the most upstream molecule in the NF-κB signal pathway. Oral administration of Ma-ME not only alleviated inflammatory lesions, but also reduced the gene expression of IL-1β and p-Syk in mice with HCl/EtOH-induced gastritis. HPLC and LC-MS/MS analyses confirmed that Ma-ME contains various anti-inflammatory flavonoids, including quercetin, daidzein, and nevadensin.
CONCLUSIONS: Ma-ME exhibited anti-inflammatory activities in vitro and in vivo by targeting Syk in the NF-κB signaling pathway. Therefore, we propose that Ma-ME could be used to treat inflammatory diseases such as gastritis.
Materials and Methods: Dried root of P. glabra was extracted under reflux with methyl alcohol, fractionated through the vacuum liquid chromatography technique, and evaporated and then purified the compounds using column chromatography and preparative thin-layer chromatography. THP-1 cells were treated with amentoflavone, 5,7,4'-hydroxyflavonoid, and stigmasterol with various concentrations (0-30 µg/mL) and then incubated with MTS reagent for 2h. Treatment was done for 24, 48, and 72h. Then, effects of these compounds were also tested on PGE2, TNF-α, and IL-6 expression in human THP-1-derived macrophage cells for 24h.
Results: Three new compounds such as amentoflavone, 5,7,4'-hydroxyflavonoid, and stigmasterol were isolated. After 24h of incubation, a significant decrease in cell viability was reported with IC50 values of amentoflavone, 5,7,4'- hydroxyflavonoid, and stigmasterol (21 µg/mL ≡ 38 M), (18 µg/mL ≡ 66 M) and (20 µg/mL ≡ 48.5 M), respectively. Whereas for 48 and 72h treatment showed a less decreased cell viability compared with 24h treatment. These compounds also showed a significant reduction in the production of TNF-α, IL-6, and PGE2 in a dose-dependent manner.
Conclusions: The isolated new compounds showed significant cytotoxicity and anti-inflammatory effects.
AIM OF THE STUDY: The present study aimed to characterize the chemical properties of a purified polysaccharide extracted from the aerial part of Tetrastigma hemsleyanum (SYQP) and investigate its antipyretic and antitumor effects in mice models.
MATERIALS AND METHODS: Water-soluble crude polysaccharides from the aerial parts of Tetrastigma hemsleyanum were extracted and fractionated by DEAE and gel permeation chromatography. Homogeneity, molecular weight, monosaccharide composition, and FTIR analysis were performed to characterize the SYQP. Antipyretic effect of SYQP was examined using Brewer's yeast induced hyperthermia test. Antitumor effect was investigated using H22 tumor bearing mice. The serum cytokines were determined to evaluated the biological activities of SYQP.
RESULTS: SYQP was composed of galacturonic acid (GalA), glucose (Glc), mannose (Man), arabinose (Ara), galactose (Gal), and rhamnose (Rha) with a molar ratio of 11.3:7.1:2.5:1.0:0.9:0.5 and it had an average molecular weight of 66.2 kDa. The oral administration of SYQP at 200 and 400 mg/kg could markedly suppress the hyperthermia of mice induced by Brewer's yeast and decrease the production of cytokines especially prostaglandin E2 (PGE2) in the serum of mice. SYQP inhibited the growth of H22 tumor in mice with inhibitory rate of 39.9% at the administration dose of 200 mg/kg and increased the production of cytokines such as tumor necrosis factor-alpha (TNF-a) and interferon γ (IFN-γ). Experimental results showed that the preventive administration of SYQP before lipopolysaccharide (LPS) reduced the high cytokine levels such as IL-6, IL-10 and IFN-γ, indicating that SYQP might act as a competitor with LPS to interact with toll like receptor 4 (TLR4), which further regulated the secretion of cytokines.
CONCLUSION: The anti-inflammatory and antitumor activities of SYQP might be related to its regulation of host immune function by controlling the secretion of cytokines.
Methods: NQC was synthesised and characterised using spectroscopic techniques. The compound was tested for its anti-inflammatory effect using Lipopolysaccharide from Escherichia coli (LPSEc) induced inflammation in mouse macrophages (RAW 264.7 cells). The effect of NQC on inflammatory cytokines was measured using enzyme-linked immune sorbent assay (ELISA). The Nrf2 activity of the compound NQC was determined using 'Keap1:Nrf2 Inhibitor Screening Assay Kit'. To obtain the insights on NQC's activity on Nrf2, molecular docking studies were performed using Schrödinger suite. The metabolic stability of NQC was determined using mouse, rat and human microsomes.
Results: NQC was found to be non-toxic at the dose of 50 µM on RAW 264.7 cells. NQC showed potent anti-inflammatory effect in an in vitro model of LPSEc stimulated murine macrophages (RAW 264.7 cells) with an IC50 value 26.13 ± 1.17 µM. NQC dose-dependently down-regulated the pro-inflammatory cytokines [interleukin (IL)-1β (13.27 ± 2.37 μM), IL-6 (10.13 ± 0.58 μM) and tumor necrosis factor (TNF)-α] (14.41 ± 1.83 μM); and inflammatory mediator, prostaglandin E2 (PGE2) with IC50 values, 15.23 ± 0.91 µM. Molecular docking studies confirmed the favourable binding of NQC at Kelch domain of Keap-1. It disrupts the Nrf2 interaction with kelch domain of keap 1 and its IC50 value was 4.21 ± 0.89 µM. The metabolic stability studies of NQC in human, rat and mouse liver microsomes revealed that it is quite stable with half-life values; 63.30 ± 1.73, 52.23 ± 0.81, 24.55 ± 1.13 min; microsomal intrinsic clearance values; 1.14 ± 0.31, 1.39 ± 0.87 and 2.96 ± 0.34 µL/min/g liver; respectively. It is observed that rat has comparable metabolic profile with human, thus, rat could be used as an in vivo model for prediction of pharmacokinetics and metabolism profiles of NQC in human.
Conclusion: NQC is a new class of NRF2 activator with potent in vitro anti-inflammatory activity and good metabolic stability.
METHODS: 2, 2'-[1, 2-cyclohexanediylbis (nitriloethylidyne)]bis(4-bromophenol) (CNBP) is synthesized via a Schiff base reaction, using the related ketone and diamine as the starting materials. SD rats are divided as normal, ulcer control (5 ml/kg of 10% Tween 20), testing (10 and 20 mg/kg of CNBP) and reference groups (omeprazole 20 mg/kg). Except for the normal group, the rest of the groups are induced gastric ulcer by ethanol 1 h after the pre-treatment. Ulcer area, gastric wall mucus, and acidity of gastric content of the animal stomachs are measured after euthanization. Antioxidant activity of the compound is tested by Ferric reducing antioxidant power (FRAP) test and safety of the compound is identified through acute toxicity by [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Moreover, activities of superoxide dismutase (SOD), catalase (CAT), levels of prostaglandins E2 (PGE2) and also malondialdehyde (MDA) are determined.
RESULTS: Antioxidant activity of CNBP was approved via FRAP assay. Vast shallow hemorrhagic injury of gastric glandular mucosa was observed in the ulcer group compared to the CNBP-treated animals. Histological evaluations confirmed stomach epithelial defense effect of CNBP with drastic decrease of gastric ulceration, edema and leucocytes penetration of submucosal stratum. Immunostaining exhibited over-expression in HSP70 protein in CNBP-treated groups compared to that of the ulcer group. Also, gastric protein analysis showed low levels of MDA, PGE2 and high activity of SOD and CAT.
CONCLUSIONS: CNBP with noticeable antioxidant property showed gastroprotective activity in the testing rodents via alteration of HSP70 protein expression. Also, antioxidant enzyme activities which were changed after treatment with CNBP in the animals could be elucidated as its gastroprotective properties.
Material and methods: The methanolic extract of PS was prepared in the dose of 500 mg/kg. Twenty-eight male Wistar rats were assigned to 4 equal sized groups: two control groups and two treated groups which were supplemented with either PS or OMZ orally at a dose of 500 mg/kg and 20 mg/kg body weight respectively. After 28 days of treatment, one control group, the PS and OMZ group were subjected to a single exposure of water-immersion restraint stress for 3.5 h. After the last exposure to stress, the stomach was excised for evaluation of the parameters.
Results: Oral supplementation of PS was as effective in preventing the formation of gastric lesion when compared with OMZ (p < 0.05). The increased gastric acidity and MDA due to stress was also reduced with supplementation of PS and OMZ. Only PS had the ability to reduce prostaglandin E2 loss (p = 0.0067) and have the ability to down regulate cyclooxygenase-2 (COX-2) mRNA expression (p = 0.01) with stress exposure.
Conclusions: Piper sarmentosum possesses a similar protective effect against stress-induced gastric lesions as omeprazole. The protective effect was associated with decreased lipid peroxidation, increased prostaglandin E2, reduction in gastric acidity and reduction in COX-2 mRNA expression which was altered by stress.