Anti-inflammatory effects of Flos Lonicerae Japonicae Water Extract are regulated by the STAT/NF-κB pathway and HO-1 expression in Virus-infected RAW264.7 cells.

This study examined the effect of the Flos Lonicerae Japonicae water extract (FLJWE), chlorogenic acid, and luteolin on pseudorabies virus (PRV)-induced inflammation in RAW264.7 cells and elucidated related molecular mechanisms. The results revealed that FLJWE and luteolin, but not chlorogenic acid, inhibited the production of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and inflammatory cytokines in PRV-infected RAW 264.7 cells. We found that the FLJWE and luteolin suppressed nuclear factor (NF)-κB activation by inhibiting the phosphorylation of signal transducer and activator of transcription 1 and 3 (STAT1 and STAT3, respectively). Moreover, the FLJWE significantly upregulated the expression of pNrf2 and its downstream target gene heme oxygenase-1 (HO-1). Our data indicated that FLJWE and luteolin reduced the expression of proinflammatory mediators and inflammatory cytokines, such as COX-2 and iNOS, through the suppression of the JAK/STAT1/3-dependent NF-κB pathway and the induction of HO-1 expression in PRV-infected RAW264.7 cells. The findings indicate that the FLJWE can be used as a potential antiviral agent.

Litchi (Litchi chinensis Sonn.) flower proanthocyanidin fraction exhibited protective efficacy to suppress nickel-induced expression for vascular endothelial growth factor in HepG2 cells.

The protective efficacy of litchi (Litchi chinensis Sonn.) flower proanthocyanidin fraction (LFPF) composed of (-)-epicatechin and proanthocyanidin A2 against vascular endothelial growth factor (VEGF) generation induced by nickel (Ni) in hepatocellular carcinoma (Hep G2) cells was studied. VEGF is an angiogenic inducer, which promotes tumor angiogenesis, leading to rapid tumor growth and metastasis. VEGF could be substantially induced in the Ni-mediated Hep G2 cells. Through LFPF treatment, the Ni-induced VEGF generation could be suppressed significantly. The inhibition of HIF-1α expression by blocking phosphatidylinositol-3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) pathways, and the suppression of Janus kinase 2 (JAK2)/signal transducers and activators of transcription 3 (STAT 3), and Raf-1 proto-oncogene, serine/threonine kinase (RAF1)/mitogen-activated protein kinase (MEK1/2)/extracellular-signal-regulated kinase (ERK1/2) pathways are important molecular mechanisms for the LFPF action. LFPF should probably reduce the risk of liver cancer in Ni-contaminated environments by inhibiting VEGF expression. PRACTICAL APPLICATIONS: LFPF mainly contained (-)-epicatechin and proanthocyanidin A2. Our results demonstrated that LFPF considerably suppressed the Ni-induced VEGF expression through inhibition of JAK2/STAT 3 and RAF1/MEK1/2/ERK1/2 pathways and prohibited HIF-1α expression through blocking PI3K/AKT/mTOR pathway. Litchi flowers might have the potential to diminish the liver cancer risk in a Ni-contaminated environment through suitable treatment.

Preventive effects of taurine against d-galactose-induced cognitive dysfunction and brain damage.

Oxidative stress arising from life processes or environmental influences and its resultant cellular dysfunctions are major causes of neurodegenerative disorders. The objectives of this study were to investigate whether taurine (Tau) can prevent d-galactose-induced cognitive dysfunction and brain oxidative damage. Mice given with Tau supplementation (100 and 400 mg per kg BW per day) spent shorter (p < 0.05) time in searching target in d-galactose (100 mg per kg BW per day) treated mice in a water maze reference memory experiment. Moreover, Tau supplementation extended (p < 0.05) the searching period around the target quadrant in the probe test of the water maze, and neuronal degeneration and nucleus shrinkage in the hippocampus dentate gyrus area of d-galactose treated mice were observed to be attenuated. Tau also downregulated (p < 0.05) expression of the glial fibrillary acidic protein (Gfap) and of the cluster of differentiation marker Cd11b; meanwhile, it strengthened (p < 0.05) antioxidant capacity and lowered (p < 0.05) the accumulation of advanced glycation end-products (AGEs) in the brain. Therefore, Tau could be effective to ameliorate oxidative damage and inflammation in the brain, and apoptosis of brain cells, which further lessen the cognitive dysfunction.

Dunaliella salina alga extract inhibits the production of interleukin-6, nitric oxide, and reactive oxygen species by regulating nuclear factor-κB/Janus kinase/signal transducer and activator of transcription in virus-infected RAW264.7 cells.

Recent investigations have demonstrated that carotenoid extract of Dunaliella salina alga (Alga) contains abundant ß-carotene and has good anti-inflammatory activities. Murine macrophage (RAW264.7 cells) was used to establish as an in vitro model of pseudorabies virus-induced reactive oxygen species (ROS) response. In this study, antioxidant activities of Alga were measured based on 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging, trolox equivalent antioxidant capacity assays, reducing power, and virus-induced ROS formation in RAW264.7 cells. Anti-inflammatory activities of Alga were assessed by its ability to inhibit the production of interleukin-6 and nitric oxide (NO) using enzyme-linked immunosorbent assay, then the Janus kinase/signal transducer and activator of transcription (JAK/STAT) pathway was investigated by measuring the inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), nuclear factor-κB (p50 and p65), JAK, STAT-1/3, and suppressor of cytokine signaling 3 (SOCS3) by Western blotting. In addition, Alga inhibited virus replication by plaque assay. Our results showed that the Alga had high antioxidant activity, significantly reduced the virus-induced accumulation of ROS, and inhibited the levels of nitric oxide and interleukin-6. Further studies revealed that Alga also downregulated the gene and protein expressions of iNOS, COX-2, nuclear factor-κB (p50 and p65), and the JAK/STAT pathway. The inhibitory effects of Alga were similar to pretreatment with specific inhibitors of JAK and STAT-3 in pseudorabies virus -infected RAW264.7 cells. Alga enhanced the expression of SOCS3 to suppress the activity of the JAK/STAT signaling pathway in pseudorabies virus-infected RAW264.7 cells. In addition, Alga has decreased viral replication (p < 0.005) at an early stage. Therefore, our results demonstrate that Alga inhibits ROS, interleukin6, and nitric oxide production via suppression of the JAK/STAT pathways and enhanced the expression of SOCS3 in virus-infected RAW264.7 cells.

Effects of antrosterol from Antrodia camphorata submerged whole broth on lipid homeostasis, antioxidation, alcohol clearance, and anti-inflammation in livers of chronic-alcohol fed mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Antrodia camphorata is a functional fungus in Taiwan and owns several pharmacological functions. Antrosterol, a bioactive constitute of sterols in edible Antrodia camphorata submerged whole broth, can protect liver from CCl4 damage via enhancing antioxidant and anti-inflammatory capacities. AIM OF THE STUDY: The aim of this study was to investigate the hepatoprotection of antrosterol (named as EK100) against alcohol consumption. MATERIALS AND METHODS: A Lieber-DeCarli regular EtOH diet (EtOH liquid diet, 5% (v/v) alcohol) was applied to induce alcoholic liver damage. Mice were randomly divided into 5 groups: (1) Control: control liquid diet; (2) EtOH: EtOH liquid diet; (3) EK100_1X: EtOH liquid diet and 1mg EK100 (Antrosterol)/Kg body weight (bw); (4) EK100_5X: EtOH liquid diet and 5mg EK100/Kg bw; (5) EK100_10X: EtOH liquid diet and 10mg EK100/Kg bw. At the end of experiment, the livers were collected for histo-pathological analyses, RNA and protein extraction, and enzymatic activities. RESULTS: Antrosterol reduced serum/liver lipids of alcohol-diet fed mice which highly related to upregulated fatty acid ß-oxidation and downregulated lipogenesis, and increased fecal lipid/bile-acid outputs. Antrosterol enhanced hepatic antioxidant capabilities in alcohol-diet fed mice while it also lowered serum alcohol level, as well as increased alcohol dehydrogenase (ADH) and catalase (CAT) activities and decreased CYP2E1 protein expression in livers of alcohol-diet fed mice. Besides, antrosterol lowered hepatic inflammation and fibrosis related gene expressions, as well as serum AST/ALT values and TNF-α/IL-1ß contents in alcohol-diet fed mice. CONCLUSION: Based on the results, hepatoprotection of antrosterol is mostly attributed to its regulations of lipid homeostasis, antioxidant capability, alcohol metabolism, and anti-inflammation.

Molecular mechanisms of the effects of the ethanolic extract of Muntingia calabura Linn. fruit on lipopolysaccharide-induced pro-inflammatory mediators in macrophages.

Four flavonoids (epicatechin, rutin, diosmin and luteolin) and 11 phenolic acids (gallic acid, gentisic acid, p-hydroxybezoic acid, vanillic acid, caffeic acid, p-coumaric acid, ferulic acid, sinapic acid, syringic acid, p-anisic acid and rosmarinic acid) were determined in the ethanolic extract of M. calabura Linn. fruit gathered in Taiwan. The extract suppressed the lipopolysaccharide-stimulated expressions of inducible nitric oxide synthase and cyclooxygenase-2 as well as the productions of nitric oxide, prostaglandin E2 and pro-inflammatory cytokines [tumour necrosis factor-α, interleukin (IL)-1ß and IL-6] in RAW264.7 macrophages. The extract modulated the inflammatory processes through inactivation of nuclear factor-κB (NF-κB), mitogen-activated protein kinases (MAPKs) p38 and c-Jun NH2-terminal kinase 1/2 (JNK1/2), and Janus kinase 2 (JAK2)/signal transducers and activators of transcription 1/3 (STAT1/3). Moreover, the activation of nuclear factor erythroid-2-related factor 2 (Nrf2) followed by inducing the production of heme oxygenase-1 (HO-1) is also related to the anti-inflammatory effect of the extract.

Luteolin inhibits viral-induced inflammatory response in RAW264.7 cells via suppression of STAT1/3 dependent NF-κB and activation of HO-1.

Luteolin is a common dietary flavonoid present in Chinese herbal medicines that has been reported to have important anti-inflammatory properties. Previous studies have shown that luteolin is an anti-inflammatory and anti-oxidative agent. In this study, the anti-virus inflammatory capacity of luteolin and its molecular mechanisms of action were analyzed. The cytotoxic effects of luteolin were assessed in the presence or absence of pseudorabies virus (PRV) via LDH and MTT assays. The results showed that luteolin (<10µM) had no toxic effects and there were tendencies toward higher cell survival. In PRV-infected RAW264.7 cells, luteolin potently inhibited the production of NO, iNOS, COX-2 and inflammatory cytokine production. Luteolin did not inhibit the phosphorylation of ERK 1/2, p38, and JNK 1/2 either. We found that PRV-induced NF-κB activation is regulated through inhibition of STAT1and STAT3 phosphorylation in response to luteolin. Additionally, luteolin caused the induction of HO-1 via upregulation of Nrf2, both of which are involved in the secretion of proinflammatory mediators. The blockade of HO-1 expression with SnPP, a HO-1 inhibitor, attenuated HO-1 induction by luteolin and thus mitigated its anti-inflammatory effects during PRV-infected RAW264.7 cells. Taken together, our data indicate that luteolin diminishes the proinflammatory mediators NO, inflammatory cytokines and the expression of their regulatory genes, iNOS and COX-2, in PRV-infected RAW264.7 cells by inhibiting STAT1/3 dependent NF-κB activation and inducing Nrf2mediated HO-1 expression.

Lychee flower extract inhibits proliferation and viral replication of HSV-1-infected corneal epithelial cells.

PURPOSE: Herpes simplex virus type I (HSV-1) is capable of causing a wide array of human ocular diseases. Herpes simplex virus keratitis (HSK)-induced cytopathogenicity together with the chronic immune-inflammatory reaction can trigger stromal scarring, thinning, and neovascularization which may lead to permanent vision impairment. Lychee flower extract (LFE) is known for its antioxidant and anti-inflammatory effects. Therefore, in this study, we investigated the mechanism of the Statens Seruminstitut rabbit corneal (SIRC) epithelial cells infected by HSV-1 and examined the antiviral capabilities of LFE. METHODS: SIRC cells were pretreated with different concentrations of LFE (0.2, 0.1, and 0.05 µg/ml) and then infected with 1 MOI of HSV-1 for 24 h. The cell viability or morphology was evaluated in this study. In addition, the supernatants and cell extracts were collected for Cell Counting Kit-8 (CCK), plaque assay, and western blotting. RESULTS: We found that HSV-1-induced cell proliferation is regulated through inhibition of the mammalian target of rapamycin (mTOR) and p70s6k phosphorylation in response to the LFE. In addition, the LFE enhanced the autophagy protein expression (Beclin-1 and light chain 3, LC3) and decreased the viral titers. CONCLUSIONS: These results showed the antiviral capabilities and the protective effects of LFE. Taken together, our data indicate that LFE has potential as an anti-HSK (herpes simplex keratitis) for HSV-1 infection.

Regulation of virus-induced inflammatory response by Dunaliella salina alga extract in macrophages.

Previous reports have suggested that many constituents within various algal samples are able to attenuate LPS-induced inflammatory effects. To date no report has been published on the regulation of virus-induced inflammatory response of Dunaliella salina carotenoid extract. In the present study, the anti-inflammatory effect of D. salina carotenoid extract on pseudorabies virus (PRV)-infected RAW 264.7 macrophages was investigated. We evaluated the anti-inflammatory effect of D. salina carotenoid extract on PRV-infected RAW 264.7 cells by measuring cell viability, cytotoxicity, production of inflammatory mediators such as NO, iNOS, COX-2, pro-inflammatory cytokines and anti-virus replication by plaque assay. We found down-regulation of the expression of the iNOS, COX-2 and pro-inflammatory genes IL-1ß, IL-6, TNF-α, and MCP-1 in a dose-dependent manner. Although there was no effect on viral replication, there were tendencies toward lower virus titer and tendencies toward higher cell survival. Most importantly, we found that inhibition of TLR9, PI3K and Akt phosphorylation plays a crucial role in the extract-mediated NF-κB regulation by modulating IKK-IκB signaling in PRV-infected RAW264.7 cells. These results indicate that D. salina carotenoid extracts inhibited inflammation by inhibition of NF-κB activation by TLR9 dependent via PI3K/Akt inactivation.

Inhibitory effect of litchi (Litchi chinensis Sonn.) flower on lipopolysaccharide-induced expression of proinflammatory mediators in RAW264.7 cells through NF-κB, ERK, and JAK2/STAT3 inactivation.

Litchi (Litchi chinensis Sonn.) flower ethanolic extract (LFEE) was found to contain five flavanoids [total amount, 102.73 ± 5.50 mg/g of dried extract (gDE)], nine phenolic acids (total amount, 60.31 ± 4.52 mg/gDE), and proanthocyanidin A2 (79.31 ± 2.95 mg/gDE). LFEE was used to evaluate the inhibitory effects on lipopolysaccharide- (LPS-) induced pro-inflammatory mediators in RAW264.7 cells. The results showed that LFEE treatment could suppress the expressions of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), the productions of nitric oxide (NO) and prostaglandin E2 (PGE2), and the secretions of pro-inflammatory cytokines [interleukin-1ß (IL-1ß), IL-6, and tumor necrosis factor α (TNF-α)] in the LPS-mediated RAW264.7 cells. The attenuation of LPS-induced inflammatory responses by LFEE was found to be closely related to the inhibition of the translocation of nuclear factor κB (NF-κB) p50/p65 subunits correlated with suppression of the activation of the inhibitor of κB kinase (IKK) α/ß and downregulation of activation of extracellular signal-regulated kinase (ERK) and Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3).

Hepatoprotection of noni juice against chronic alcohol consumption: lipid homeostasis, antioxidation, alcohol clearance, and anti-inflammation.

Chronic alcohol consumption leads to steatohepatitis and cirrhosis. Naturally fermented noni juice (NJ) contains polyphenols, polysaccharides, and some trace minerals. This study explored protective effects of NJ against chronic alcohol consumption. Mice were assigned randomly to one of the following groups: (1) control, control liquid diet and distilled water; (2) alcohol, alcohol liquid diet and distilled water; (3) Alc+NJ_1X, alcohol liquid diet and 5 mL NJ/kg BW; (4) Alc+NJ_2X, alcohol liquid diet and 10 mL NJ/kg BW; (5) Alc+NJ_3X, alcohol and 15 mL NJ/kg BW for 4 weeks. NJ decreased (p < 0.05) serum AST, ALT, and alcohol levels and liver lipids, as well as increased (p < 0.05) daily fecal lipid outputs in alcohol-diet fed mice. NJ supplementation not only down-regulated (p < 0.05) lipogenesis but also up-regulated (p < 0.05) fatty acid ß-oxidation in livers of alcohol-diet fed mice. NJ also accelerated alcohol clearance via increased (p < 0.05) hepatic ADH and ALDH activities. NJ increased (p < 0.05) hepatic TEAC and GSH levels but decreased (p < 0.05) TBARS value and TLR2/4, P38, ERK 1/2, NFκB P65, iNOS, COX-2, TNF-α, and IL-1ß expressions in alcohol-diet fed mice. NJ promotes hepatoprotection against alcohol-induced injury due to regulations of lipid homeostasis, antioxidant status, alcohol metabolism, and anti-inflammatory responses.

Beneficial effects of noni (Morinda citrifolia L.) juice on livers of high-fat dietary hamsters.

Polyphenols in noni juice (NJ) are mainly composed of phenolic acids, mainly gentisic, p-hydroxybenoic, and chlorogenic acids. To investigate the beneficial effects of NJ on the liver, hamsters were fed with two diets, normal-fat and high-fat diets. Furthermore, high-fat dietary hamsters were received distilled water, and 3, 6, and 9 mL NJ/kg BW, respectively. After a 6-week feeding period, the increased (p<0.05) sizes of liver and visceral fat in high-fat dietary hamsters compared to the control hamsters were ameliorated (p<0.05) by NJ supplementation. NJ also decreased (p<0.05) serum/liver lipids but enhanced (p<0.05) daily faecal lipid/bile acid outputs in the high-fat dietary hamsters. High-fat dietary hamsters supplemented with NJ had higher (p<0.05) liver antioxidant capacities but lowered (p<0.05) liver iNOS, COX-2, TNF-α, and IL-1ß expressions, gelatinolytic levels of MMP9, and serum ALT values compared to those without NJ. Hence, NJ protects liver against a high-fat dietary habit via regulations of antioxidative and anti-inflammatory responses.

Effects of Antrodia camphorata on alcohol clearance and antifibrosis in livers of rats continuously fed alcohol.

Alcoholic fatty liver disease (AFLD) is the result of an excessive or chronic consumption of alcohol. Nine male Wistar rats per group were randomly assigned to one of the following drinking treatments: a 20% (w/w) alcohol solution (ALC); a 20% (w/w) alcohol solution cotreated with 0.25 g silymarin/kg BW/day; or a 20% (w/w) alcohol solution cotreated with 0.025 g Niuchangchih ( Antrodia camphorata )/kg BW/day for 4 weeks. Rats with cotreatments of silymarin or Niuchangchih had smaller (p < 0.05) relative liver size, less (p < 0.05) liver lipid accumulation, and lower (p < 0.05) liver damage indices [aspartate aminotransferase (AST) and alkaline phosphatase (ALP) values]. In the regulation of alcohol metabolism, the lower serum alcohol level was observed only in alcohol-fed rats supplemented with Niuchangchih. Meanwhile, cotreatment of silymarin or Niuchangchih increased (p < 0.05) CAT and ALDH activities but did not (p > 0.05) affect ADH and CYP2E1 expressions, which accelerate alcohol metabolism in the body. Additionally, neither silymarin nor Niuchangchih (p > 0.05) influenced serum/hepatic MMP-2 activities and NF-κB, AP1, and α-SMA gene expressions, but serum/hepatic MMP-9 activities and TNF-α, KLF-6, and TGF-ß1 gene expressions of alcohol-fed rats were down-regulated (p < 0.05) by silymarin or Niuchangchih, which also could explain the lower liver damage observed in rats chronically fed alcohol.

Antiobesity and hypolipidemic effects of polyphenol-rich longan (Dimocarpus longans Lour.) flower water extract in hypercaloric-dietary rats.

Plenty of polyphenols, i.e. phenolic acids and flavonoids, were found in longan flower water extract (LFWE) through spectrophotometric and HPLC analyses. Antiobesity and hypolipidemic effects of polyphenol-rich longan flower water extract (LFWE) were investigated in this study. Eight male rats per group were assigned randomly to one of the following dietary groups: (1) normal-caloric diet and pure water (NCD + NDW); (2) hypercaloric diet and pure water (HCD + NDW); (3) HCD and 1.25% (w/v) LFWE (HCD + 1.25% LFWE); (4) HCD and 2.5% (w/v) LFWE (HCD + 2.5% LFWE) for 9 weeks. Body weight, size of epididymal fat, serum triglyceride level and atherogenic index, and hepatic lipids were decreased (p < 0.05) in HCD rats by drinking 2.5% LFWE which may result from downregulated (p < 0.05) pancreatic lipase activity, and sterol regulatory element binding protein-1c (SREBP-1c) and fatty acid synthase (FAS) gene expressions, as well as upregulated (p < 0.05) LDL receptor (LDLR) and peroxisome proliferator-activated-receptor-alpha (PPAR-alpha) gene expressions, and also increased (p < 0.05) fecal triglyceride excretions. Therefore, polyphenol-rich LFWE indeed characterizes antiobesity and hypolipidemic effects in vivo.

Flaxseed oil attenuates nonalcoholic fatty liver of hyperlipidemic hamsters.

Hyperlipidemia of hamsters was induced by high-fat/cholesterol diets formulated by the addition of coconut oil (CO), butter (BU), and flaxseed oil (FX). Lower (p < 0.05) serum lipids, liver size, and hepatic cholesterol and triacylglycerol contents were observed in the FX group compared to both CO and BU groups. The liver damage indices [glutamic oxaloacetic transaminase (GOT) and glutamic pyruvic transaminase (GPT) values] in the FX group were lower (p < 0.05) than those in the CO and BU groups, which may result from higher (p < 0.05) glutathione (GSH) levels and a tendency toward lower malondialdehyde (MDA) levels in livers. Besides, lower (p < 0.05) gene expression and activity of hepatic matrix metalloproteinases-9 (MMP-9) in the FX group were lower (p > 0.05) compared to those in the CO and BU groups; however, no (p > 0.05) differences in gene expression activities of hepatic MMP-2 were observed among treatments. Those beneficial effects could explain the attenuation of FX on nonalcoholic fatty liver (NAFL) induced by a high-fat/cholesterol dietary habit.