Anti-Neuroinflammatory and Neuroprotective Effect of Intermedin B Isolated from the Curcuma longa L. via NF-κB and ROS Inhibition in BV2 Microglia and HT22 Hippocampal Cells.

Compounds derived from Curcuma longa L. (C. longa) have been extensively studied and reported to be effective and safe for the prevention and treatment of various diseases, but most research has been focused on curcuminoids derived from C. longa. As neurodegenerative diseases are associated with oxidation and inflammation, the present study aimed to isolate and identify active compounds other than curcuminoids from C. longa to develop substances to treat these diseases. Seventeen known compounds, including curcuminoids, were chromatographically isolated from the methanol extracts of C. longa, and their chemical structures were identified using 1D and 2D NMR spectroscopy. Among the isolated compounds, intermedin B exhibited the best antioxidant effect in the hippocampus and anti-inflammatory effect in microglia. Furthermore, intermedin B was confirmed to inhibit the nuclear translocation of NF-κB p-65 and IκBα, exerting anti-inflammatory effects and inhibiting the generation of reactive oxygen species, exerting neuroprotective effects. These results highlight the research value of active components other than curcuminoids in C. longa-derived compounds and suggest that intermedin B may be a promising candidate for the prevention of neurodegenerative diseases.

Antioxidant Capacity of Potentilla paradoxa Nutt. and Its Beneficial Effects Related to Anti-Aging in HaCaT and B16F10 Cells.

Skin aging is a natural process influenced by intrinsic and extrinsic factors, and many skin anti-aging strategies have been developed. Plants from the genus Potentilla has been used in Europe and Asia to treat various diseases. Potentilla paradoxa Nutt. has been used as a traditional medicinal herb in China and has recently been shown to have anti-inflammatory effects. Despite the biological and pharmacological potential of Potentilla paradoxa Nutt., its skin anti-aging effects remain unclear. Therefore, this study evaluated the free radical scavenging, moisturizing, anti-melanogenic, and wound-healing effects of an ethanol extract of Potentilla paradoxa Nutt. (Pp-EE). Pp-EE was found to contain phenolics and flavonoids and exhibits in vitro antioxidant activities. α-Linolenic acid was found to be a major component of Pp-EE on gas chromatography-mass spectrometry. Pp-EE promoted the expression of hyaluronic acid (HA) synthesis-related enzymes and suppressed the expression of HA degradation-related enzymes in keratinocytes, so it may increase skin hydration. Pp-EE also showed inhibitory effects on the production and secretion of melanin in melanocytes. In a scratch assay, Pp-EE improved skin wound healing. Taken together, Pp-EE has several effects that may delay skin aging, suggesting its potential benefits as a natural ingredient in cosmetic or pharmaceutical products.

Cardamine komarovii flower extract reduces lipopolysaccharide-induced acute lung injury by inhibiting MyD88/TRIF signaling pathways.

In the present study, we investigated anti-inflammatory effect of Cardamine komarovii flower (CKF) on lipopolysaccharide (LPS)-induced acute lung injury (ALI). We determined the effect of CKF methanolic extracts on LPS-induced pro-inflammatory mediators NO and prostaglandin E2 (PGE2), production of pro-inflammatory cytokines (IL-1ß, TNF-α, and IL-6), and related protein expression levels of MyD88/TRIF signaling pathways in peritoneal macrophages (PMs). Nuclear translocation of NF-κB-p65 was analyzed by immunofluorescence. For the in vivo experiments, an ALI model was established to detect the number of inflammatory cells and inflammatory factors (IL-1ß, TNF-α, and IL-6) in bronchoalveolar lavage fluid (BALF) of mice. The pathological damage in lung tissues was evaluated through H&E staining. Our results showed that CKF can decrease the production of inflammatory mediators, such as NO and PGE2, by inhibiting their synthesis-related enzymes iNOS and COX-2 in LPS-induced PMs. In addition, CKF can downregulate the mRNA levels of IL-1ß, TNF-α, and IL-6 to inhibit the production of inflammatory factors. Mechanism studies indicated that CKF possesses a fine anti-inflammatory effect by regulating MyD88/TRIF dependent signaling pathways. Immunocytochemistry staining showed that the CKF extract attenuates the LPS-induced translocation of NF-kB p65 subunit in the nucleus from the cytoplasm. In vivo experiments revealed that the number of inflammatory cells and IL-1ß in BALF of mice decrease after CKF treatment. Histopathological observation of lung tissues showed that CKF can remarkably improve alveolar clearance and infiltration of interstitial and alveolar cells after LPS stimulation. In conclusion, our results suggest that CKF inhibits LPS-induced inflammatory response by inhibiting the MyD88/TRIF signaling pathways, thereby protecting mice from LPS-induced ALI.

Anti-inflammatory action of Athyrium multidentatum extract suppresses the LPS-induced TLR4 signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: The aerial part of Athyrium multidentatum (Doll.) Ching (AM) is widely used in the northeastern region of China as an edible wild herb, but its medicinal value, especially its anti-inflammatory effect, has not been fully explored. AIM OF THE STUDY: To investigate the anti-inflammatory activity of AM and clarify the anti-inflammatory mechanism involving the TLR4 signaling pathway using a lipopolysaccharide (LPS)-induced inflammatory model. MATERIALS AND METHODS: AM ethanol extract was used as the experimental material to investigate the effect that the extract has on the production of pro-inflammatory mediators (NO, PGE2, TNF-α, IL-1ß and IL-6); changes in LPS-induced peritoneal macrophages (PMs); and TLR4-mediated intracellular events, including MAPKs (ERK, JNK, and p38) and IκB-α in the MyD88-dependant pathway and IRF3, STAT1, and STAT3 in the TRIF-dependent pathway. In in vivo experiments, we established an LPS-induced acute lung injury (ALI) model and investigated the cell count and cytokine (TNF-α, IL-1ß and IL-6) levels in bronchoalvelar lavage fluid (BALF) of C57BL6 mice. Histological changes in the lung tissues were observed with H&E staining. RESULTS: AM extract inhibited NO and PGE2 by suppressing their synthetase (iNOS and COX-2) gene expression in LPS-induced PMs; the secretion of IL-6, IL-1ß, and TNF-α also deceased via the down-regulation of mRNA levels. Furthermore, the TLR4-mediated intracellular events involved the phosphorylated forms of MAPKs (ERK, JNK) and IκB-α in the MyD88-dependent pathway and the TRIF-dependent pathway (IRF3, STAT1, STAT3), and the relevant proteins were expressed at low levels in the AM extract groups. In in vivo experiments, the cell count and cytokine (TNF-α, IL-1ß and IL-6) levels in BALF decreased significantly in a dose-dependent manner in the AM extract groups. The lung tissue structure exhibited dramatic damage in the LPS group, and the damaged area decreased in the AM extract groups; in particular, the effect of 10 mg/kg extract was similar to that of the positive control dexamethasone (DEX). CONCLUSION: The findings demonstrate that AM protects against LPS-induced acute lung injury by suppressing TLR4 signaling, provide scientific evidence to support further study of the safety of anti-inflammatory drugs and indicate that AM can be used as an anti-inflammatory and anti-injury agent to prevent pneumonia caused by microbial infection.

Taraxacum coreanum protects against glutamate-induced neurotoxicity through heme oxygenase-1 expression in mouse hippocampal HT22 cells.

Taraxacum coreanum Nakai is a dandelion that is native to Korea, and is widely used as an edible and medicinal herb. The present study revealed the neuroprotective effect of this plant against glutamate-induced oxidative stress in HT22 murine hippocampal neuronal cells. Ethanolic extracts from the aerial (TCAE) and the root parts (TCRE) of T. coreanum were prepared. Both extracts were demonstrated, by high performance liquid chromatography, to contain caffeic acid and ferulic acid as representative constituents. TCAE and TCRE significantly increased cell viability against glutamate-induced oxidative stress in mouse hippocampal HT22 cells. Western blot analysis revealed that treatment of HT22 cells with the extracts induced increased expression of the enzyme heme oxygenase-1 (HO-1), compared with untreated cells, in a concentration-dependent manner. Increased HO-1 enzymatic activity, compared with untreated cells, was also demonstrated following treatment with TCAE and TCRE. In addition, western blot analysis of the nuclear fractions of both TCAE and TCRE-treated HT22 cells revealed increased levels of nuclear factor erythroid 2 like 2 (Nrf2) compared with untreated cells, and decreased Nrf2 levels in the cytoplasmic fraction compared with untreated cells. The present study suggested that the neuroprotective effect of T. coreanum is associated with induction of HO-1 expression and Nrf2 translocation to the nucleus. Therefore, T. coreanum exhibits a promising function in prevention of neurodegeneration. Further studies will be required for the isolation and the full characterization of its active substances.

Cearoin Induces Autophagy, ERK Activation and Apoptosis via ROS Generation in SH-SY5Y Neuroblastoma Cells.

Neuroblastomas are the most common solid extracranial tumors in childhood. We investigated the anticancer effect of cearoin isolated from Dalbergia odorifera in human neuroblastoma SH-SY5Y cells. SH-SY5Y cells were treated with various doses of cearoin. The viability was measured by MTT assay. DCFDA fluorescence assay and Griess assay were used for the measurement of intracellular reactive oxygen species (ROS) and nitric oxide (NO), respectively. Western blot analysis was performed to clarify the molecular pathway involved. Cearoin induced cell death in a dose-dependent manner. Cearoin increased the phosporylation of ERK, the conversion of LC3B-I to LC3B-II, decrease in Bcl2 expression, the activation of caspase-3, and the cleavage of PARP, indicating the induction of autophagy and apoptosis. Furthermore, cearoin treatment increased the production of ROS and NO. Co-treatment with the antioxidant N-acetylcysteine completely abolished cearoin-mediated autophagy, ERK activation and apoptosis, suggesting the critical role of ROS in cearoin-induced anticancer effects. Moreover, co-treatment with ERK inhibitor PD98059 partially reversed cearoin-induced cell death, indicating the involvement of ERK in cearoin anticancer effects. These data reveal that cearoin induces autophagy, ERK activation and apoptosis in neuroblastoma SH-SY5Y cells, which is mediated primarily by ROS generation, suggesting its therapeutic application for the treatment of neuroblastomas.

Rhamnella gilgitica Attenuates Inflammatory Responses in LPS-Induced Murine Macrophages and Complete Freund's Adjuvant-Induced Arthritis Rats.

Rhamnella gilgitica Mansf. et Melch, which belongs to the rhamnus family (Rhamnaceae), is traditionally used to treat rheumatism, swelling and pain in China. However, little is known about the pharmacological activities of this plant. The anti-inflammatory activities of the 70% ethanol extract of R. gilgitica (RG) in RAW264.7 macrophages and complete Freund's adjuvant (CFA)-induced arthritic rats are investigated in this study for the first time. The effects of RG on cell viability were determined by a MTT assay, and the effects of RG on pro-inflammatory mediators were analyzed by ELISA and Western blot. The effects of RG on paw thickness, thymus and spleen index were also examined in CFA-induced arthritic rats. RG suppressed the induction of proinflammatory mediators, including iNOS (inducible nitric oxide synthase), NO (nitric oxide), COX-2 (cyclooxygenase-2) and PG (prostaglandin) E2 in LPS stimulated RAW264.7 macrophages. RG also inhibited the phosphorylation and degradation of I[Formula: see text]B-[Formula: see text], as well as the nuclear translocation of nuclear factor kappa B (NF-[Formula: see text]B) p65. In addition, RG treatment significantly reduced the paw thickness in CFA-induced arthritic rats. Oral administration of RG led to a significant decrease of both the thymus and spleen index at a concentration of 100[Formula: see text]mg/mL. Taken together, these findings suggest that RG might be an agent for further development in the treatment of a variety of inflammatory diseases.

Protective effect of ganodermanondiol isolated from the Lingzhi mushroom against tert-butyl hydroperoxide-induced hepatotoxicity through Nrf2-mediated antioxidant enzymes.

Ganodermanondiol, a biologically active compound, was isolated from the Lingzhi mushroom (Ganoderma lucidum). The present study examined the protective effects of ganodermanondiol against tert-butyl hydroperoxide (t-BHP)-induced hepatotoxicity. Ganodermanondiol protected human liver-derived HepG2 cells through nuclear factor-E2-related factor 2 (Nrf2) pathway-dependent heme oxygenase-1 expressions. Moreover, ganodermanondiol increased cellular glutathione levels and the expression of the glutamine-cysteine ligase gene in a dose-dependent manner. Furthermore, ganodermanondiol exposure enhanced the phosphorylation of adenosine monophosphate-activated protein kinase (AMPK) and its upstream kinase activators, LKB1 and Ca(2+)/calmodulin-dependent protein kinase-II (CaMKII). This study indicates that ganodermanondiol exhibits potent cytoprotective effects on t-BHP-induced hepatotoxicity in human liver-derived HepG2 cells, presumably through Nrf2-mediated antioxidant enzymes and AMPK.

Involvement of Heme Oxygenase-1 Induction in the Cytoprotective and Immunomodulatory Activities of Viola patrinii in Murine Hippocampal and Microglia Cells.

A number of diseases that lead to injury of the central nervous system are caused by oxidative stress and inflammation in the brain. In this study, NNMBS275, consisting of the ethanol extract of Viola patrinii, showed potent antioxidative and anti-inflammatory activity in murine hippocampal HT22 cells and BV2 microglia. NNMBS275 increased cellular resistance to oxidative injury caused by glutamate-induced neurotoxicity and reactive oxygen species generation in HT22 cells. In addition, the anti-inflammatory effects of NNMBS275 were demonstrated by the suppression of proinflammatory mediators, including proinflammatory enzymes (inducible nitric oxide synthase and cyclooxygenase-2) and cytokines (tumor necrosis factor-α and interleukin-1ß). Furthermore, we found that the neuroprotective and anti-inflammatory effects of NNMBS275 were linked to the upregulation of nuclear transcription factor-E2-related factor 2-dependent expression of heme oxygenase-1 in HT22 and BV2 cells. These results suggest that NNMBS275 possesses therapeutic potential against neurodegenerative diseases that are induced by oxidative stress and neuroinflammation.

Involvement of heme oxygenase-1 induction in inhibitory effect of ethyl gallate isolated from Galla Rhois on nitric oxide production in RAW 264.7 macrophages.

In the present study, we investigated an anti-inflammatory effect of ethyl gallate (EG) isolated from Galla Rhois as evaluated by inhibition of nitric oxide (NO) production and inducible nitric oxide synthase (iNOS) expression, and a potential role of heme oxygenase-1 (HO-1) in the inhibition of NO production elicited by EG. Treatment of RAW264.7 macrophages with EG significantly inhibited the production of NO and iNOS expression stimulated by lipopolysaccharide (LPS). We also demonstrated that EG treatment increased HO-1 mRNA and protein expression, as assessed by quantitative RT-PCR and Western blot analysis. EG treatment also increased the levels of nuclear factor-erythroid 2-related factor 2, which is critical for transcriptional induction of HO-1. In addition, treatment with SnPP (tin protoporphyrin IX), a selective HO-1 inhibitor, counteracted the inhibitory effect of EG on nitrite production, suggesting that HO-1 is, at least in part, implicated in the inhibition of NO production induced by EG treatment. Taken together, these results indicate that EG isolated from Galla Rhois suppresses NO production in LPS-stimulated RAW 264.7 macrophages via HO-1 induction.

Neuroprotective effects of constituents of the root bark of Dictamnus dasycarpus in mouse hippocampal cells.

Glutamate-induced oxidative injury causes neuronal degeneration related to many central nervous system diseases, such as Parkinson's disease, Alzheimer's disease, epilepsy and ischemia. The bioassay-guided fractionation of the EtOH extract of the root bark of Dictamnus dasycarpus Trucz. provided one neuroprotective limonoid, obacunone, together with a degraded limonoid, fraxinellone and two alkaloids, dictamine and haplopine. At concentrations of 100-150 microM, obacunone showed the potent neuroprotective effects on glutamateinduced neurotoxicity and induced the expression of heme oxygenase (HO)-1 in the mouse hippocampal HT22 cells. In addition, we found that obacunone increased p38 MAPK phosphorylation and induced HO-1 expression via p38 MAPK pathway. These results suggest that obacunone isolated from the root bark of D. dasycarpus increases cellular resistance to glutamate-induced oxidative injury in mouse hippocampal HT22 cells, presumably through the p38 MAPK pathway-dependent HO-1 expression. These results suggest that obacunone could be the effective candidates for the treatment of ROS-related neurological diseases.

Tribuli fructus constituents protect against tacrine-induced cytotoxicity in HepG2 cells.

A new phenolic amide, tribulusimide D (4-hydroxy-N-[3-(4-hydroxy-3-methoxyphenyl)-1-oxo-2-propen-1-yl]-3-methoxybenzamide) (1), together with a known phenolic amide, terrestriamide ((E)-3-(4-hydroxy-3-methoxyphenyl)-N-[2-(4-hydroxyphenyl)-2-oxoethyl]-prop-2-enamide) (2) and a flavonol glycoside, quercetin-3-O-beta-D-glucopyranosyl-(1-->6)-beta-D-glucopyranoside (3) were isolated from the H2O extract of Tribuli Fructus. Compounds 1 and 3 showed significant hepatoprotective activities, with EC50 values of 13.46 +/- 0.2 and 7.06 +/- 0.7 microM, respectively, against tacrine-induced cytotoxicity in HepG2 cells.

Chromone glycosides and hepatoprotective constituents of Hypericum erectum.

Two chromone glycosides, hyperimone A [7-(beta-D-glucopyranosyloxy)-5-hydroxy-2-(1-methylethyl)-4H-1-benzopyran-4-one (1)] and hyperimone B [7-(beta-D-glucopyranosyloxy)-5-hydroxy-3-methyl-4H-1-benzopyran-4-one (2)], together with six known compounds were isolated from the methanolic extract of the whole plant of Hypericum erectum. 1,3,5,6-Tetrahydroxyxanthone (5) and I3, II8-biapigenin (6) showed moderate hepatoprotective activity with EC(50) values of 160.2 +/- 0.6 microM and 217.7 +/- 1.3 microM, respectively, against tacrine-induced cytotoxicity in HepG2 cells.

Cytoprotective constituents of the heartwood of Caesalpinia sappan on glutamate-induced oxidative damage in HT22 cells.

The bioassay-guided fractionation of a MeOH extract of the heartwood of Caesalpinia sappan L. provided two neuroprotective compounds, sappanchalcone (2) and 4-O-methylepisappanol (3), together with a methoxychalcone, isoliquiritigenin 2'-methyl ether (1), and three aromatic compounds, 4-O-methylsappanol (4), caesalpine J (5), pluchoic acid (6). At concentrations of 20-40 microM, compound 2 showed significant cytoprotective effects against glutamate-induced oxidative stress through the induction of heme oxygenase (HO)-1 in HT22-immortalized hippocampal cells. Compound 3 also showed moderate neuroprotective effect at 40 microM, but compounds 1, 4-6 did not show any protective effects against glutamate-induced cytotoxicity in HT22 cells.

Flavonoids from the heartwood of Dalbergia odorifera and their protective effect on glutamate-induced oxidative injury in HT22 cells.

Two flavonoids, 4,2',5'-trihydroxy-4'-methoxychalcone (1) and (2S)-6,7,4'-trihydroxyflavan (2), along with fourteen known flavonoids and two other known arylbenzofurans were isolated from the heartwood of Dalbergia odorifera. The structure of compounds 1 and 2 were established by spectroscopic (NMR and MS) analyses. Of the isolates, eight compounds (1, 4, 7, 10, 12, 14, 15, 17) were found to have potent protective effect on glutamate-induced oxidative injury in HT22 cells.

Cudratricusxanthone A protects mouse hippocampal cells against glutamate-induced neurotoxicity via the induction of heme oxygenase-1.

Cudratricusxantone A (CTXA), isolated from the roots of Cudrania tricuspidata Bureau (Moraceae), has potent hepatoprotective, antiproliferative, and monoamine oxidase inhibitory effects. In this study, we examined whether CTXA could protect HT22-immortalized hippocampal cells against glutamate-induced oxidative stress through the induction of heme oxygenase (HO)-1 expression. CTXA induced the expression of HO-1 and increased HO activity dose- and time-dependently. CTXA also suppressed glutamate-induced ROS generation in HT22 cells. Interestingly, treatment of neuronal cells with CTXA enhanced cellular resistance to glutamate oxidative stress. The protective effect of CTXA was abrogated by tin protoporphyrin IX, an HO inhibitor. In addition, treatment with the HO-1 inducer, cobalt protoporphyrin IX, and bilirubin, one of the enzymatic products of HO-1, produced comparable protection. These results demonstrate that CTXA protects neuronal cells from glutamate-induced oxidative stress via the induction of HO-1.

1,2,3,4,6-penta-O-galloyl-beta-D-glucose from Galla Rhois protects primary rat hepatocytes from necrosis and apoptosis.

In this study, we investigated the hepatoprotective effects of four compounds from Galla Rhois [gallic acid methyl ester, gallic acid, an equilibrium mixture of 3-galloyl-gallic acid and 4-galloyl-gallic acid isomers, and 1,2,3,4,6-penta- O-galloyl- beta- D-glucose (PGG)] in primary rat hepatocytes undergoing necrosis or apoptosis. Treatment with gallic acid methyl ester (12.5 and 50 microM) or PGG (3.125, 12.5 and 50 microM) reduced hepatocyte necrosis induced by tert-butyl hydroperoxide. PGG treatment (4 and 20 microM) also altered hepatocyte apoptosis induced by glycochenodeoxycholic acid. Based on these results, we propose that PGG warrants further evaluation as a hepatoprotective agent, because it protected primary rat hepatocytes from both necrosis and apoptosis.

In vitro hepatoprotective compounds from Suaeda glauca.

Bioassay-guided fractionation of the MeOH extract of Suaeda glauca yielded four phenolic compounds, methyl 3,5-di-O-caffeoyl quinate (1) and 3,5-di-O-caffeoyl quinic acid (2), isorhamnetin 3-O-beta-D-galactoside (3), and quercetin 3-O-beta-D-galactoside (4). Compounds 1 and 2 were hepatoprotective against tacrine-induced cytotoxicity in human liver-derived Hep G2 cells with the EC(50) values of 72.7+/-6.2 and 117.2+/-10.5 microM, respectively. Silybin as a positive control showed an EC(50) value of 82.4+/-4.1 microM.

Heme oxygenase-1 inducing constituent of Prunella vulgaris in HepG2 cells.

Inducible heme oxygenase (HO)-1 is known to play a major role in the pathogenesis of several diseases, and it protects cells against oxidant-mediated injury. The bioassay-guided fractionation of the EtOH extract of the flowered fruit-spike of Prunella vulgaris L. (Labiatae) yielded two ursane-type triterpenes, 3beta,23-dihydroxyurs-12-en-28-oic acid (23-hydroxyursolic acid) (1) and 3beta-hydroxyurs-12-en-28-oic acid (ursolic acid) (2). Western blotting demonstrated that treatment with compound 1 increased the expression of HO-1 in a dose-dependent manner in human liver-derived HepG2 cells. Investigation of structure-related HO-1 inducing activity suggested that the hydroxyl group at the C-23 position in the ursane skeleton is important for this activity.

Cytoprotective constituent of Hoveniae Lignum on both Hep G2 cells and rat primary hepatocytes.

A phytochemical investigation of the EtOH extract of Hoveniae Lignum yielded four phenolic compounds, phloretin (1), 5-(4'-hydroxyphenyl)-gamma-valerolactone (2), (-)-epiafzelechin (3), and maesopsin (4). Compound 1 was hepatoprotective against tacrine-induced cytotoxicity in human liver-derived Hep G2 cells with an EC50 value of 37.55 +/- 0.42 microM. Compound 1 (0.4-200 microM) also significantly reduced tert-butyl hydroperoxide-induced cytotoxicity in rat primary hepatocytes as measured by the cellular leakage of lactate dehydrogenase and the level of aspartate transaminase.