Hongming Lv

Xanthohumol ameliorates lipopolysaccharide (LPS)-induced acute lung injury via induction of AMPK/GSK3β-Nrf2 signal axis

Abundant natural flavonoids can induce nuclear factor-erythroid 2 related factor 2 (Nrf2) and/or AMP-activated protein kinase (AMPK) activation, which play crucial roles in the amelioration of various inflammation- and oxidative stress-induced diseases, including acute lung injury (ALI). Xanthohumol (Xn), a principal prenylflavonoid, possesses anti-inflammation and anti-oxidant activities. However, whether Xn could protect from LPS-induced ALI through inducing AMPK/Nrf2 activation and its downstream signals, are still poorly elucidated. Accordingly, we focused on exploring the protective effect of Xn in the context of ALI and the involvement of underlying molecular mechanisms. Our findings indicated that Xn effectively alleviated lung injury by reduction of lung W/D ratio and protein levels, neutrophil infiltration, MDA and MPO formation, and SOD and GSH depletion. Meanwhile, Xn significantly lessened histopathological changes, reactive oxygen species (ROS) generation, several cytokines secretion, and iNOS and HMGB1 expression, and inhibited Txnip/NLRP3 inflammasome and NF-κB signaling pathway activation. Additionally, Xn evidently decreased t-BHP-stimulated cell apoptosis, ROS generation and GSH depletion but increased various anti-oxidative enzymes expression regulated by Keap1-Nrf2/ARE activation, which may be associated with AMPK and GSK3β phosphorylation. However, Xn-mediated inflammatory cytokines and ROS production, histopathological changes, Txnip/NLRP3 inflammasome and NF-κB signaling pathway in WT mice were remarkably abrogated in Nrf2-/- mice. Our experimental results firstly provided a support that Xn effectively protected LPS-induced ALI against oxidative stress and inflammation damage which are largely dependent upon upregulation of the Nrf2 pathway via activation of AMPK/GSK3β, thereby suppressing LPS-activated Txnip/NLRP3 inflammasome and NF-κB signaling pathway.

Isovitexin Exerts Anti-Inflammatory and Anti-Oxidant Activities on Lipopolysaccharide-Induced Acute Lung Injury by Inhibiting MAPK and NF-κB and Activating HO-1/Nrf2 Pathways

Oxidative damage and inflammation are closely associated with the pathogenesis of acute lung injury (ALI). Thus, we explored the protective effect of isovitexin (IV), a glycosylflavonoid, in the context of ALI. To accomplish this, we created in vitro and in vivo models by respectively exposing macrophages to lipopolysaccharide (LPS) and using LPS to induce ALI in mice. In vitro, our results showed that IV treatment reduced LPS-induced pro-inflammatory cytokine secretion, iNOS and COX-2 expression and decreased the generation of ROS. Consistent findings were obtained in vivo. Additionally, IV inhibited H2O2-induced cytotoxicity and apoptosis. However, these effects were partially reversed following the use of an HO-1 inhibitor in vitro. Further studies revealed that IV significantly inhibited MAPK phosphorylation, reduced NF-κB nuclear translocation, and upregulated nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase 1 (HO-1) expression in RAW 264.7 cells. In vivo, pretreatment with IV attenuated histopathological changes, infiltration of polymorphonuclear granulocytes and endothelial activation, decreased the expression of ICAM-1 and VCAM-1, reduced the levels of MPO and MDA, and increased the content of GSH and SOD in ALI. Furthermore, IV treatment effectively increased Nrf2 and HO-1 expression in lung tissues. Therefore, IV may offer a protective role against LPS-induced ALI by inhibiting MAPK and NF-κB and activating HO-1/Nrf2 pathways.

Tenuigenin ameliorates acute lung injury by inhibiting NF-κB and MAPK signalling pathways.

We aimed to explore the protective effect of tenuigenin (TNG) on lipopolysaccharide (LPS)-stimulated inflammatory responses in acute lung injury (ALI). Thus, we assessed the effects of TNG on the LPS-induced production of tumour necrosis factor (TNF)-α, interleukin (IL)-6, and IL-1β in the culture supernatants of RAW 264.7 cells. Male BALB/c mice were pretreated with commercial TNG (2, 4 and 8 mg/kg) and dexamethasone (Dex, 5mg/kg) for 1h prior to LPS (0.5 mg/kg) challenge. After 12h, airway inflammation was assessed. Our results showed that TNG dramatically decreased the production of TNF-α, IL-1β, and IL-6 in vitro and in vivo as well as the expression of COX-2 protein in vivo. Treatment with TNG not only significantly ameliorated LPS-stimulated histopathological changes but also reduced the myeloperoxidase (MPO) activity and the wet-to-dry weight ratio of the lungs. Furthermore, TNG blocked IκBα phosphorylation and degradation and inhibited p38/ERK phosphorylation in LPS-induced ALI. These findings suggest that TNG may have a protective effect on LPS-induced ALI and may be useful for the prevention and treatment of ALI in the clinical setting.

Nrf2-mediated liver protection by esculentoside A against acetaminophen toxicity through the AMPK/Akt/GSK3β pathway.

Acetaminophen (APAP) overdose accounts for the majority of acute liver failure cases, and oxidative stress plays a key role in its toxic effects. Esculentoside A (EsA) has anti-oxidant activities, but its therapeutic potential for APAP hepatotoxicity remains unknown. This study aimed to assess the protective effects and mechanism of EsA against APAP-induced hepatotoxicity in vitro and in vivo. In vitro, EsA treatment inhibited APAP- or H2O2-induced cytotoxicity, H2O2 and O2- production, glutathione (GSH) depletion and apoptosis dependent on nuclear factor erythroid-2-related factor 2 (Nrf2) activation in HepG2 cells. Moreover, EsA significantly increased the phosphorylation of AMP-activated protein kinase (AMPK) and serine/threonine kinase (Akt), as well as glycogen synthase kinase 3 beta (GSK-3β) inhibitory phosphorylation at Ser9. Furthermore, an AMPK inhibitor (compound c) abolished the effects of EsA on AKT phosphorylation, GSK-3β inactivation, Nrf2 nuclear translocation and cytoprotection. With regard to APAP-induced acute liver injury, EsA attenuated the APAP-stimulated increases in the serum ALT and AST levels, as well as centrilobular necrosis and GSH depletion in the mice. In addition, it decreased the GSSG level, GSSG-to-GSH ratio, and the phosphorylation and mitochondrial translocation of c-Jun N-terminal kinase (JNK). Further, the protective potential of EsA against mitochondrial dysfunction was exhibited not only by inhibiting Bax mitochondrial translocation and the release of mitochondrial inter-membrane proteins, such as apoptosis-inducing factor (AIF), but also by activating Nrf2/HO-1. Collectively, our findings suggest that EsA has protective potential against APAP toxicity by potentiating the Nrf2-regulated survival mechanism through the AMPK/Akt/GSK3β pathway.

Daphnetin-mediated Nrf2 antioxidant signaling pathways ameliorate tert-butyl hydroperoxide (t-BHP)-induced mitochondrial dysfunction and cell death

Abstract Daphnetin (Daph), a natural coumarin derivative isolated from plants of the Genus Daphne, possesses abundant biological activities, such as anti‐inflammatory, antioxidant and anticancer properties. In the present study, we focused on investigating the protective effect of Daph against tert‐butyl hydroperoxide (t‐BHP)‐induced oxidative damage, mitochondrial dysfunction and the involvement of underlying molecular mechanisms. Our findings indicated that Daph effectively inhibited t‐BHP‐stimulated cytotoxicity, cell apoptosis, and mitochondrial dysfunction, which are associated with suppressed reactive oxygen species (ROS) generation, decreased malondialdehyde (MDA) formation, increased superoxide dismutase (SOD) levels and glutathione (GSH)/GSSG (oxidized GSH) ratio. Further investigation indicated that Daph significantly suppressed cytochrome c release and NLRP3 inflammasome activation and modulated apoptosis‐related protein Bcl‐2, Bax, and caspase‐3 expression. Moreover, Daph dramatically induced the expression of the glutamate‐cysteine ligase modifier (GCLM) subunit and the glutamate‐cysteine ligase catalytic (GCLC) subunit, heme oxygenase‐1 (HO‐1), and NAD (P) H: quinone oxidoreductase (NQO1), which is largely dependent on upregulating the nuclear factor‐erythroid 2‐related factor 2 (Nrf2) nuclear translocation, reducing the Keap1 protein expression, and strengthening the antioxidant response element (ARE) promoter activity. Additionally, Daph remarkably activated a c‐Jun NH2‐terminal kinase (JNK) and extracellular signal‐regulated kinase (ERK) phosphorylation, but ERK and JNK inhibitor pretreatment exhibited an evident decrease of the level of Daph‐enhanced Nrf2 nuclear translocation. Furthermore, Daph exposure suppressed t‐BHP‐induced cytotoxicity and ROS overproduction, which are mostly blocked in Nrf2 knockout RAW 264.7 cells and peritoneal macrophages. Accordingly, Daph exhibited protective roles against t‐BHP‐triggered oxidative damage and mitochondrial dysfunction by the upregulation of Nrf2 antioxidant signaling pathways, which may be involved in the activation of JNK and ERK. Graphical abstract Scheme summarizing the protection of t‐BHP‐induced RAW 264.7 cell mitochondrial dysfunction by Daph via activation of the Keap1‐Nrf2/ARE signaling pathway. Figure. No Caption available. HighlightsDaph inhibits t‐BHP‐induced oxidative damage and mitochondrial dysfunction.Daph blocks cytochrome c release and NLRP3 inflammasome activation.Daph modulates apoptosis‐related protein Bcl‐2, Bax, and caspase‐3 expression.Daph induces expressions of GCLC, GCLM, HO‐1 and NQO‐1.Daph upregulates Keap‐1‐Nrf2/ARE signaling pathways via activation of JNK and ERK.

Hesperetin Suppresses Inflammatory Responses in Lipopolysaccharide-Induced RAW 264.7 Cells via the Inhibition of NF-κB and Activation of Nrf2/HO-1 Pathways

Hesperetin (Hesp), a common flavanone glycoside, was extracted from the fruit peel of Citrus aurantium L. (Rutaceae). Hesp has been shown to possess various biological properties, including antioxidant, neuroprotective, and anti-inflammatory properties. In this study, we investigated the protective effect of Hesp on inflammatory responses in lipopolysaccharide (LPS)-induced RAW 264.7 cells. Our results indicated that Hesp treatment dramatically suppressed secretion of tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-1β; reduced inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) gene expression; inhibited NF-κB (p65) phosphorylation; and blocked IκBα phosphorylation and degradation. Further studies revealed Hesp markedly enhanced the heme oxygenase (HO)-1 and nuclear factor erythroid 2-related factor 2 (Nrf2) expression, which were involved with inducing Nrf2 nuclear translocation and decreasing Keap1 protein expression. Together, these results indicated that the anti-inflammatory effect of Hesp may be associated with NF-κB inhibition and Nrf2/HO-1 activation.

Lico A Enhances Nrf2-Mediated Defense Mechanisms against t-BHP-Induced Oxidative Stress and Cell Death via Akt and ERK Activation in RAW 264.7 Cells

Licochalcone A (Lico A) exhibits various biological properties, including anti-inflammatory and antioxidant activities. In this study, we investigated the antioxidative potential and mechanisms of Lico A against tert-butyl hydroperoxide- (t-BHP-) induced oxidative damage in RAW 264.7 cells. Our results indicated that Lico A significantly inhibited t-BHP-induced cytotoxicity, apoptosis, and reactive oxygen species (ROS) generation and reduced glutathione (GSH) depletion but increased the glutamate-cysteine ligase modifier (GCLM) subunit and the glutamate-cysteine ligase catalytic (GCLC) subunit genes expression. Additionally, Lico A dramatically upregulated the antioxidant enzyme heme oxygenase 1 (HO-1) and nuclear factor erythroid 2-related factor 2 (Nrf2), which were associated with inducing Nrf2 nuclear translocation, decreasing Keap1 protein expression and increasing antioxidant response element (ARE) promoter activity. Lico A also obviously induced the activation of serine/threonine kinase (Akt) and extracellular signal-regulated kinase (ERK), but PI3K/Akt and ERK inhibitors treatment displayed clearly decreased levels of LicoA-induced Nrf2 nuclear translocation and HO-1 expression, respectively. Furthermore, Lico A treatment markedly attenuated t-BHP-induced oxidative damage, which was reduced by treatment with PI3K/Akt, ERK, and HO-1 inhibitors. Therefore, Lico A might have a protective role against t-BHP-induced cytotoxicity by modulating HO-1 and by scavenging ROS via the activation of the PI3K/Akt and ERK/Nrf2 signaling pathways.

D(-)-Salicin inhibits the LPS-induced inflammation in RAW264.7 cells and mouse models.

D(-)-Salicin is a traditional medicine which has been known to exhibit anti-inflammation and other therapeutic activities. The present study aimed to investigate whether D(-)-Salicin inhibited the LPS-induced inflammation in vivo and in vitro. We evaluated the effect of D(-)-Salicin on cytokines (TNF-α, IL-1β, IL-6 and IL-10) in vivo and in vitro by enzyme-linked immunosorbent assay and signaling pathways (MAPKs and NF-κB) in vivo by Western blot. The results showed that D(-)-Salicin markedly decreased TNF-α, IL-1β and IL-6 concentrations and increased IL-10 concentration. In addition, western blot analysis indicated that D(-)-Salicin suppressed the activation of MAPKs and NF-κB signaling pathways stimulated by LPS. To examine whether D(-)-Salicin ameliorated LPS-induced lung inflammation, inhibitors of MAPKs and NF-κB signaling pathways were administrated intraperitoneally to mice. Interference with specific inhibitors revealed that D(-)-Salicin-mediated cytokine suppression was through MAPKs and NF-κB pathways. In the mouse model of acute lung injury, histopathologic examination indicted that D(-)-Salicin suppressed edema induced by LPS. So it is suggest that D(-)-Salicin might be a potential therapeutic agent against inflammatory diseases.

Asiatic Acid Exhibits Anti-inflammatory and Antioxidant Activities against Lipopolysaccharide and d-Galactosamine-Induced Fulminant Hepatic Failure

Inflammation and oxidative stress are essential for the pathogenesis of fulminant hepatic failure (FHF). Asiatic acid (AA), which is a pentacyclic triterpene that widely occurs in various vegetables and fruits, has been reported to possess antioxidant and anti-inflammatory properties. In this study, we investigated the protective effects of AA against lipopolysaccharide (LPS) and d-galactosamine (GalN)-induced FHF and the underlying molecular mechanisms. Our findings suggested that AA treatment effectively protected against LPS/d-GalN-induced FHF by lessening the lethality; decreasing the alanine transaminase and aspartate aminotransferase levels, interleukin (IL)-1β, IL-6, and tumor necrosis factor-α production, malondialdehyde formation, myeloperoxidase level and reactive oxygen species generation (i.e., H2O2, NO, and O2−), and increasing the glutathione and superoxide dismutase contents. Moreover, AA treatment significantly inhibited mitogen-activated protein kinase (MAPK) and nuclear factor-kappa B (NF-κB) signaling pathway activation via the partial induction of programmed cell death 4 (PDCD4) protein expressions, which are involved in inflammatory responses. Furthermore, AA treatment dramatically induced the expression of the glutamate-cysteine ligase modifier subunit, the glutamate-cysteine ligase catalytic subunit, heme oxygenase-1, and NAD (P) H: quinoneoxidoreductase 1 (NQO1), which are largely dependent on activation of the nuclear factor-erythroid 2-related factor 2 (Nrf2) through the induction of AMP-activated protein kinase (AMPK) and glycogen synthase kinase-3β (GSK3β) phosphorylation. Accordingly, AA exhibited protective roles against LPS/d-GalN-induced FHF by inhibiting oxidative stress and inflammation. The underlying mechanism may be associated with the inhibition of MAPK and NF-κB activation via the partial induction of PDCD4 and upregulation of Nrf2 in an AMPK/GSK3β pathway activation-dependent manner.

Betulin exhibits anti-inflammatory activity in LPS-stimulated macrophages and endotoxin-shocked mice through an AMPK/AKT/Nrf2-dependent mechanism

Continued oxidative stress can lead to chronic inflammation, which in turn could mediate most chronic diseases including cancer. Nuclear factor erythroid 2-related factor (Nrf2), a critical transcriptional activator for antioxidative responses, has envolved to be an attractive drug target for the treatment or prevention of human diseases. In the present study, we investigated the effects and mechanisms of betulin on Nrf2 activation and its involvement in the lipopolysaccharide (LPS)-triggered inflammatory system. In macrophages, betulin activated the Nrf2 signaling pathway and increased Nrf2-targeted antioxidant and detoxifying enzymes, including NADPH, quinine oxidoreductase 1 (NQO1), heme oxygenase-1 (HO-1), γ-glutamyl cysteine synthetase catalytic subunit (GCLC) and modifier subunit (GCLM) in a dose and time dependent manner. Importantly, we found betulin-induced activation of Nrf2 is AMPK/AKT/GSK3β dependent, as pharmacologically inactivating AMPK blocked the activating effect of betulin on AKT, GSK3β and Nrf2. Furthermore, betulin attenuated LPS-induced inflammatory mediators (iNOS and COX-2) and MAPK inflammatory signaling pathway. The effect of betulin on HO-1 and NQO1 upregulation, iNOS and COX-2 the downregulation, and survival time extension was largely weakened when Nrf2 was depleted in vitro and in vivo. Our results demonstrate that the AMPK/AKT/Nrf2 pathways are essential for the anti-inflammatory effects of betulin in LPS-stimulated macrophages and endotoxin-shocked mice.