Palbinone from Paeonia suffruticosa Protects Hepatic Cells via Up‐regulation of Heme Oxygenase‐1

Paeonia suffruticosa has been traditionally employed for vitalizing blood circulation and alleviating liver and inflammatory diseases. The pathways by which palbinone (PB) isolated from P. suffruticosa mediates heme oxygenase‐1 (HO‐1) induction were investigated using the specific inhibitors for PI3K and mitogen activated protein kinases pathways. The effect of PB‐treatment on Nrf2 translocalization and HO‐1‐antioxidant response element (ARE) regulation was examined employing Western blot and luciferase assays. PB induced HO‐1 expression via the activation of Nrf2 in the hepatic cells, and ARE‐dependent genes were stimulated via the PB‐mediated Nrf2 activation. PB‐mediated HO‐1 expression could be involved with PI3K/Akt and ERK1/2 pathways. Our study suggests the mechanism by which PB induces HO‐1 expression in the hepatic cells. This might substantiate the traditional applications of P. suffruticosa for the treatment of oxidative stress‐related diseases including oxidant and inflammatory‐mediated vascular and liver diseases. Copyright © 2013 John Wiley & Sons, Ltd.     

Eupatolide Inhibits PDGF‐induced Proliferation and Migration of Aortic Smooth Muscle Cells Through ROS‐dependent Heme Oxygenase‐1 Induction

The abnormal proliferation and migration of vascular smooth muscle cell (VSMC) contributes importantly to the pathogenesis of atherosclerosis and restenosis. Here, we investigated the effects of eupatolide (EuTL), a sesquiterpene lactone isolated from the medicinal plant Inula britannica, on platelet‐derived growth factor (PDGF)‐induced proliferation and migration of primary rat aortic smooth muscle cells (RASMCs), as well as its underlying mechanisms. EuTL remarkably inhibited PDGF‐induced proliferation and migration of RASMCs. Treatment of RASMCs with EuTL induced both protein and mRNA expression of heme oxygenase‐1 (HO‐1). SB203580 (a p38 inhibitor), SP600125 (a JNK inhibitor), U0126 (a MEK inhibitor) and LY294002 (a PI3K inhibitor) did not suppress EuTL‐induced HO‐1 expression; however, N‐acetylcysteine (NAC, an antioxidant) blocked EuTL‐induced HO‐1 expression. Moreover, treatment of RASMCs with EuTL increased reactive oxygen species (ROS) accumulation and nuclear translocation of nuclear factor‐E2‐related factor 2 (Nrf2); however, this translocation was also inhibited by NAC. NAC or inhibition of HO‐1 significantly attenuated the inhibitory effects of EuTL on PDGF‐induced proliferation and migration of RASMCs. Taken together, these findings suggest that EuTL could suppress PDGF‐induced proliferation and migration of VSMCs through HO‐1 induction via ROS‐Nrf2 pathway and may be a potential HO‐1 inducer for preventing or treating vascular diseases. Copyright © 2013 John Wiley & Sons, Ltd.     

Sour Cherry Seed Kernel Extract Increases Heme Oxygenase‐1 Expression and Decreases Representation of CD3+ TNF‐α + and CD3 + IL‐8+ Subpopulations in Peripheral Blood Leukocyte Cultures from Type 2 Diabetes Patients

The present study evaluates a hypothesis that sour cherry (Prunus cerasus) seed extracts (SCE) modulate CD3+ T lymphocyte activity in ways predictive of potential for uses of SCE in management of inflammatory diseases. Peripheral blood mononuclear cells (PBMC) from 12 type 2 diabetes (T2DM) patients and eight healthy control subjects were cultured 24 h with 100 ng/ml lipopolysaccharide (LPS) to increase inflammatory signaling and co‐incubated with 0.5–100 µg/ml SCE. Cultures were evaluated by two‐color flow cytometry for percent representation of CD3+ IL8+ and CD3 + TNF‐α + cells which express interleukin‐8 (IL‐8), and tumor necrosis factor‐α, (TNF‐α+) respectively, and by enzyme‐linked immunoassay for lymphocyte‐associated heme oxygenase‐1 (HO‐1, known to be induced by SCE). SCE dosage ranges of 0.5–100 µg/ml in cell cultures significantly suppressed LPS‐increased CD3 + TNF‐α + and CD3 + IL8+ representation from all participants (p < 0.05), with greater pharmacological effect noted in suppression of CD3 + TNF‐α + noted in cells from T2DM patients versus healthy control subjects. These effects correlated with increased HO‐1 expression in SCE‐treated PBMC from all subjects (p < 0.05). Since TNF‐α and IL‐8 are diagnostic/prognostic biomarkers for many inflammatory syndromes, the capacity of SCE to down‐regulate representation of cells that express them suggests potential for therapeutic use of SCE in T2DM and other diseases. Copyright © 2012 John Wiley & Sons, Ltd.     

Sanguisorbae Radix Protects Against 6‐Hydroxydopamine‐induced Neurotoxicity by Regulating NADPH Oxidase and NF‐E2‐related Factor‐2/Heme Oxygenase‐1 Expressions

6‐Hydroxydopamine (6‐OHDA) produces neuronal cell damage by generating reactive oxygen species (ROS). The major mechanisms of protection against ROS‐induced stress are inhibiting expression of ROS generating genes such as NADPH oxidase (NOX) and increasing expression of endogenous antioxidant genes such as heme oxygenase‐1 (HO‐1). This study investigated whether a standardized Sanguisorbae Radix extract (SRE), a medical herb commonly used in Asian traditional medicine, has a protective effect on 6‐OHDA‐induced cell toxicity by regulating ROS in SH‐SY5Y cells. SRE at 10 and 50 µg/mL significantly reduced 6‐OHDA‐induced cell damage dose dependently in the 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide assay and by Hoechst 33342 staining. SRE increased the B‐cell lymphoma 2 (Bcl‐2)/Bcl‐2‐associated X ratio and decreased cytochrome C release and caspase‐3 activity. SRE also abolished 6‐OHDA‐induced ROS by inhibiting NOX expression and by inducing HO‐1 expression via NF‐E2‐related factor‐2 activation. Taken together, these results demonstrate that SRE has protective effects against 6‐OHDA‐induced cell death by regulating ROS in SH‐SY5Y cells. Copyright © 2012 John Wiley & Sons, Ltd.     

Anti‐inflammatory properties of cryptolepine

Cryptolepine is the major alkaloid of the West African shrub, Cryptolepis sanguinolenta. Cryptolepine has been shown to inhibit nitric oxide production, and DNA binding of Nuclear Factor‐kappa B following inflammatory stimuli in vitro. In order to validate the anti‐inflammatory property of this compound in vivo, we investigated its effects on a number of animal models of inflammation. Cryptolepine (10–40 mg/kg i.p.) produced significant dose‐dependent inhibition of the carrageenan‐induced rat paw oedema, and carrageenan‐induced pleurisy in rats. These effects were compared with those of the non‐steroidal anti‐inflammatory drug indomethacin (10 mg/kg). At doses of 10–40 mg/kg i.p., cryptolepine inhibited lipopolysaccharide (LPS)‐induced microvascular permeability in mice in a dose‐related fashion. Oral administration of up to 40 mg/kg of the compound for four consecutive days did not induce gastric lesion formation in rats. Analgesic activity was also exhibited by cryptolepine through a dose‐related (10–40 mg/kg i.p.) inhibition of writhing induced by i.p. administration of acetic acid in mice. The results of this study reveal that cryptolepine possesses in vivo anti‐inflammatory activity. Copyright © 2009 John Wiley & Sons, Ltd.     

Caffeoylserotonin Protects Human Keratinocyte HaCaT Cells against H2O2‐Induced Oxidative Stress and Apoptosis through Upregulation of HO‐1 Expression via Activation of the PI3K/Akt/Nrf2 Pathway

Caffeoylserotonin (CaS) has strong radical scavenging activity as well as antioxidant activities, protecting cells from lipid peroxidation, intracellular reactive oxygen species generation, DNA damage, and cell death. The molecular mechanism by which CaS protects against oxidative stress is not well understood. Here, we analyzed the cytoprotective activity of CaS in hydrogen peroxide (H₂O₂)‐treated keratinocyte HaCaT cells. H₂O₂ induced apoptosis in the cells through activation of pro‐apoptotic p21, Bax, and caspase‐3. Pretreatment with CaS inhibited apoptotic gene expression and activated the anti‐apoptotic gene, Bcl‐xL. Although CaS did not directly affect heme oxygenase‐1 (HO‐1) expression, pretreatment with CaS augmented HO‐1 expression through an increase in NF‐E2‐related factor (Nrf2) stability and stimulation of Nrf2 translocation to the nucleus upon H₂O₂ exposure. H₂O₂ also induced the phosphorylation and subsequent activation of ERK, p38 MAPK, and Akt. Analysis using specific inhibitors of p38 MAPK and Akt demonstrated that only Akt activation was involved in HO‐1 and Nrf2 expressions. In addition, PI3K and PKC inhibitors suppressed HO‐1/Nrf2 expression and Akt phosphorylation. These results demonstrate that CaS protects against oxidative stress‐induced keratinocyte cell death in part through the activation of Nrf2‐mediated HO‐1 induction via the PI3K/Akt and/or PKC pathways, but not MAPK signaling. Copyright © 2013 John Wiley & Sons, Ltd.     

Asiaticoside Inhibits TNF‐α‐Induced Endothelial Hyperpermeability of Human Aortic Endothelial Cells

The increase in endothelial permeability often promotes edema formation in various pathological conditions. Tumor necrosis factor‐alpha (TNF‐α), a pro‐atherogenic cytokine, impairs endothelial barrier function and causes endothelial dysfunction in early stage of atherosclerosis. Asiaticoside, one of the triterpenoids derived from Centella asiatica, is known to possess antiinflammatory activity. In order to examine the role of asiaticoside in preserving the endothelial barrier, we assessed its effects on endothelial hyperpermeability and disruption of actin filaments evoked by TNF‐α in human aortic endothelial cells (HAEC). TNF‐α caused an increase in endothelial permeability to fluorescein isothiocyanate (FITC)‐dextran. Asiaticoside pretreatment significantly suppressed TNF‐α‐induced increased permeability. Asiaticoside also prevented TNF‐α‐induced actin redistribution by suppressing stress fiber formation. However, the increased F to G actin ratio stimulated by TNF‐α was not changed by asiaticoside. Cytochalasin D, an actin depolymerizing agent, was used to correlate the anti‐hyperpermeability effect of asiaticoside with actin cytoskeleton. Surprisingly, asiaticoside failed to prevent cytochalasin D‐induced increased permeability. These results suggest that asiaticoside protects against the disruption of endothelial barrier and actin rearrangement triggered by TNF‐α without a significant change in total actin pool. However, asiaticoside seems to work by other mechanisms to maintain the integrity of endothelial barrier rather than stabilizing the F‐actin organization. Copyright © 2015 John Wiley & Sons, Ltd.     

Anti‐inflammatory Effects of Ginsenosides Rg5, Rz1, and Rk1: Inhibition of TNF‐α‐induced NF‐κB,COX‐2, and iNOS transcriptional expression

In the course of this experiment on the anti‐inflammatory effect of ginsenosides, protopanaxdiol ginsenosides have shown inhibition activities in inflammatory responses: NF‐κB, COX‐2, and iNOS were induced by TNF‐α. The responses of this experiment were evaluated by NF‐κB‐luciferase assay and RT‐PCR experiment of COX‐2 and iNOS genes. The NF‐κB expressions were inhibited by ginsenosides Rd, Rg₅, Rz₁, and Rk₁ in a dose‐dependent manner. The IC₅₀ values were 3.47, 0.61, 0.63, and 0.75 μM, respectively. Particularly, ginsenosides Rg₅, Rz₁, and Rk₁ as converted ginsenosides from primary protopanaxdiol ginsenosidess significantly inhibited COX‐2 and iNOS gene expression. These inhibition levels were similar to sulfasalazine as reference material. Copyright © 2014 John Wiley & Sons, Ltd.     

Houttuynia cordata Thunb. Volatile Oil Exhibited Anti‐inflammatory Effects In Vivo and Inhibited Nitric Oxide and Tumor Necrosis Factor‐α Production in LPS‐stimulated Mouse Peritoneal Macrophages In Vitro

Houttuynia cordata Thunb. (HC) is a medicinal herb that generally used in traditional Chinese medicine for treating allergic inflammation. The present study investigated the inhibitory effect of the volatile oil from HC Thunb. on animal models of inflammation and the production of inflammatory mediators in vivo and in vitro. In vivo, xylene‐induced mouse ear edema, formaldehyde‐induced paw edema and carrageenan‐induced mice paw edema were significantly decreased by HC volatile oil. HC volatile oil showed pronounced inhibition of prostaglandin (PG) E₂ and malondialdehyde production in the edematous exudates. In vitro exposure of mouse resident peritoneal macrophages to 1, 10, 100 and 1000 µg/mL of HC volatile oil significantly suppressed lipopolysaccharide (LPS)‐stimulated production of NO and tumor necrosis factor‐α (TNF‐α) in a dose‐dependent manner. Exposure to HC volatile oil had no effect on cell viability and systemic toxicity. Furthermore, HC volatile oil inhibited the production of NO and TNF‐α by down‐regulating LPS‐stimulated iNOS and TNF‐α mRNA expression. Western blot analysis showed that HC volatile oil attenuated LPS‐stimulated synthesis of iNOS and TNF‐α protein in the macrophages, in parallel. These findings add a novel aspect to the biological profile of HC and clarify its anti‐inflammatory mechanism. Copyright © 2012 John Wiley & Sons, Ltd.     

Anti‐inflammatory Activity of Constituents Isolated from Aerial Part of Angelica acutiloba Kitagawa

Recently, the resources of medicinal plants have been exhausting. The root of Angelica acutiloba is one of the most important ingredients in Japanese Kampo medicine for the treatment of gynecological diseases. In our search for alternative medicinal plant resources of the root of A. acutiloba, we found that its aerial part has the anti‐inflammatory potency as well as the root. Phytochemical investigation of the aerial part resulted in the isolation of four compounds including a new dimeric phthalide, namely tokiaerialide (2), along with Z‐ligustilide (1), falcarindiol (3), and bergaptol (4). Next, we investigated the in vitro anti‐inflammatory activity of 1–4 in lipopolysaccharide‐stimulated RAW264 macrophages. Among the isolated compounds, 1 exhibited the most potent inhibition against lipopolysaccharide‐induced production of prostaglandin E₂, nitric oxide, and pro‐inflammatory cytokines (interleukin‐6 and tumor necrosis factor‐α). Compounds 3 and 4 also inhibited all inflammatory mediators, but their inhibitory abilities were weaker than those of 1. Furthermore, 1, 3, and 4 strongly also induced heme oxygenase‐1. These results suggest that 1, 3, and 4 potentially exert anti‐inflammatory activity, and the aerial part of A. acutiloba may be considered to be a useful medicinal resource for inflammatory diseases. Copyright © 2015 John Wiley & Sons, Ltd.     

Anti‐inflammatory Activities of Eleven Centaurea Species Occurring in the Carpathian Basin

Our study aimed at the identification of anti‐inflammatory activities of different fractions of C. sadleriana extract after per os administration in rats, the identification of the active compounds of the plant and the investigation of the in vitro anti‐inflammatory activities of Centaurea species native to or cultivated in the Carpathian Basin. The aerial parts of Centaurea sadleriana Janka have been used in Hungarian folk medicine to treat the wounds of sheep. Methanol extract of C. sadleriana was fractioned by solvent‐solvent partitioning. The n‐hexane fraction was further fractionated and the anti‐inflammatory activities of certain subfractions were confirmed in vivo in rats. The n‐hexane and chloroform fraction of the methanol extract of C. sadleriana exhibited remarkable COX‐1 and COX‐2 inhibiting effects in vitro. Chromatographic separation of the fractions led to the identification of the active subfractions and 11 compounds (α‐linolenic acid, γ‐linolenic acid, stigmasterol, β‐sitosterol, campesterol, vanillin, pectolinarigenin, salvigenin, hispidulin, chrysoeriol and apigenin). The in vitro screening for anti‐inflammatory activities of further Centaurea species occurring in the Carpathian Basin (C. adjarica, C. bracteata, C. cataonica, C. cynaroides, C. dealbata, C. indurata, C. macrocephala, C. melitensis, C. nigrescens, C. ruthenica) revealed considerable COX‐1 and COX‐2 inhibitory activities. Because C. sadleriana is an endangered species native only to the Carpathian Basin, the investigation of more prevalent species is reasonable. Copyright © 2012 John Wiley & Sons, Ltd.     

Brassinin,a phytoalexin in cruciferous vegetables,suppresses obesity‐induced inflammatory responses through the Nrf2‐HO‐1 signaling pathway in an adipocyte‐macrophage co‐culture system

The aim of this study was to investigate the effect of brassinin (BR), a phytoalexin found in plants belonging to the Brassicaceae family, on the obesity‐induced inflammatory response and its molecular mechanism in co‐culture of 3T3‐L1 adipocytes and RAW264.7 macrophages. BR effectively suppressed lipid accumulation by down‐regulating the expression of adipogenic factors, which in turn, were regulated by early adipogenic factors such as CCAAT‐enhancer‐binding protein‐β and Kruppel‐like factor 2. Production of inflammatory cytokines and reactive oxygen species, induced by adipocyte‐conditioned medium, was significantly decreased in BR‐treated cells. This effect of BR was more prominent in contact co‐culture of adipocytes and macrophages with a 90% and 34% reduction in IL‐6 and MCP‐1 levels, respectively. BR also restored adiponectin expression, which was significantly reduced by culturing adipocytes in macrophage‐conditioned medium. In the transwell system, BR increased the protein levels of nuclear factor (erythroid‐derived 2)‐like 2 (Nrf2) and its target molecule, hemoxygenase‐1 (HO‐1), by 55%–93% and 45%–48%, respectively, and also increased Nrf2 translocation into the nucleus. However, knockdown of Nrf2 or HO‐1 in RAW264.7 cells restored this BR‐mediated inhibition of IL‐6 and MCP‐1 production. These results indicated that BR inhibited obesity‐induced inflammation via the Nrf2‐HO‐1 pathway.     

Anti‐inflammatory effects of chemical components from Ginkgo biloba L. male flowers on lipopolysaccharide‐stimulated RAW264.7 macrophages

Ginkgo biloba L., well known as living fossil, have various pharmacological activities. Eighteen compounds were isolated from Ginkgo male flowers including a novel matsutake alcohol glycoside, Ginkgoside A ( 1 ), and 17 known compounds—calaliukiuenoside ( 2 ), benzylalcohol O‐α‐l ‐arabinopyranosyl‐(1 → 6)‐β‐d ‐glucopyranoside ( 3 ), amentoflavone ( 4 ), sciadopitysin ( 5 ), bilobetin ( 6 ), isoginkgetin ( 7 ), olivil 4‐O‐β‐d ‐glucopyranoside ( 8 ), dihydrodehydrodiconiferyl alcohol‐4‐O‐β‐d ‐glucoside ( 9 ), (+)‐cyclo‐olivil‐6‐O‐β‐d ‐glucopyranoside ( 10 ), (?)‐isolariciresinol 4‐O‐β‐d ‐glucopyranoside ( 11 ), coniferin ( 12 ), trans‐cinnamic acid‐4‐O‐β‐d ‐glucopyranoside ( 13 ), p‐coumaryl alchol glucoside ( 14 ), stroside B ( 15 ), methylconiferin ( 16 ), cis‐p‐coumaric acid 4‐O‐β‐d ‐glucopyranoside ( 17 ), and cis‐coniferin ( 18 ). Thirteen of these compounds had not previously found in Ginkgo. All extractive fractions and isolated compounds were evaluated for their anti‐inflammatory ability in the lipopolysaccharide‐induced RAW264.7 macrophages. The ethanol extract of Ginkgo flowers and the chloroform and ethyl acetate fractions can significantly decrease nitric oxide (NO), interleukin‐6 (IL‐6), and prostaglandin E₂ (PGE₂) production at 100 μg/ml. The most effective compounds, bilobetin ( 6 ) and isoginkgetin ( 7 ), elevated the NO inhibition ratios to 80.19% and 82.37% at 50 μM, respectively. They also exhibited significant dose‐dependent inhibitory effects on tumor necrosis factor‐α, IL‐6, PGE₂, inducible NO synthase mRNA, and cyclooxygenase‐2 mRNA levels. So they can be promising candidates for the development of new anti‐inflammatory agents.