Su Jung Hwang

Anti-inflammatory effects of chlorogenic acid in lipopolysaccharide-stimulated RAW 264.7 cells

Objectives and designChlorogenic acid, which belongs to the polyphenols, is an anti-oxidant and anti-obesity agent. In this study, we investigated the role of chlorogenic acid in inflammation.Materials and methodsAnti-inflammatory effects of chlorogenic acid were examined in lipopolysaccharide (LPS)-stimulated murine RAW 264.7 macrophages and BV2 microglial cells. We observed the level of various inflammation markers such as nitric oxide (NO), inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2), tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6), and chemokine (C-X-C motif) ligand 1 (CXCL1) under LPS treatment with or without chlorogenic acid. To clarify the specific effect of chlorogenic acid, we evaluated the adhesion activity of macrophages and ninjurin1 (Ninj1) expression level in macrophages. Finally, we confirmed the activation of the nuclear factor-κB (NF-κB) signaling pathway, which is one of the most important transcription factors in the inflammatory process.ResultsChlorogenic acid significantly inhibited not only NO production but also the expression of COX-2 and iNOS, without any cytotoxicity. Chlorogenic acid also attenuated pro-inflammatory cytokines (including IL-1β and TNF-α) and other inflammation-related markers such as IL-6 in a dose-dependent manner. Additionally, endotoxin-induced adhesion of macrophages and the expression level of ninjurin1 (Ninj1) were decreased by chlorogenic acid. Finally, chlorogenic acid inhibited the nuclear translocation of NF-κB.ConclusionsChlorogenic acid may be beneficial for the prevention and treatment of anti-inflammatory diseases.

Chlorogenic acid inhibits hypoxia-induced angiogenesis via down-regulation of the HIF-1α/AKT pathway

BackgroundThe hypoxia-inducible factor-1 (HIF-1) is known to play an important role in cellular responses to hypoxia, including the transcriptional activation of a number of genes involved in tumor angiogenesis. Chlorogenic acid (CGA), one of the most abundant polyphenols in the human diet, has been reported to inhibit cancer cell growth. The effect of CGA on tumor angiogenesis and its underlying mechanisms are, as yet, unknown.MethodsThe effect of CGA on HIF-1α expression was assessed by Western blot and reverse transcriptase-polymerase chain reaction (RT-PCR) assays in A549 lung cancer cells. The transcriptional activity of the HIF-1 complex was confirmed using a luciferase assay. To assess whether angiogenic factors are increased under hypoxic conditions in these cells, vascular endothelial growth factor (VEGF) expression levels were measured by RT-PCR and Western blotting. The direct effect of CGA on human vascular endothelial cells (HUVEC) under hypoxic conditions was analyzed using in vitro assays, including tube-formation, wound healing and Transwell invasion assays. To investigate the effect of CGA on angiogenesis in vivo, we performed a Matrigel plug assay in a mouse model. Finally, the effect of CGA on AKT and ERK activation (phosphorylation) as a putative mechanism underlying the effect of CGA on VEGF-mediated angiogenesis inhibition was assessed using Western blotting.ResultsWe found that CGA significantly decreases the hypoxia-induced HIF-1α protein level in A549 cells, without changing its mRNA level. CGA was, however, found to suppress the transcriptional activity of HIF-1α under hypoxic conditions, leading to a decrease in the expression of its downstream target VEGF. We also found that CGA can block hypoxia-stimulated angiogenesis in vitro and VEGF-stimulated angiogenesis in vivo using HUVEC cells. In addition, we found that CGA can inhibit the HIF-1α/AKT signaling pathway, which plays an important role in VEGF activation and angiogenesis.ConclusionsOur data indicate that CGA plays a role in the suppression of angiogenesis via inhibition of the HIF-1α/AKT pathway. CGA may represent a novel therapeutic option for the treatment of (lung) cancer.

Green synthesis of gold nanoparticles using chlorogenic acid and their enhanced performance for inflammation.

UNLABELLED Here we developed a novel green synthesis method for gold nanoparticles (CGA-AuNPs) using chlorogenic acid (CGA) as reductants without the use of other chemicals and validated the anti-inflammatory efficacy of CGA-AuNPs in vitro and in vivo. The resulting CGA-AuNPs appeared predominantly spherical in shape with an average diameter of 22.25±4.78nm. The crystalline nature of the CGA-AuNPs was confirmed by high-resolution X-ray diffraction and by selected-area electron diffraction analyses. High-resolution liquid chromatography/electrospray ionization mass spectrometry revealed that the caffeic acid moiety of CGA forms quinone structure through a two-electron oxidation causing the reduction of Au(3+) to Au(0). When compared to CGA, CGA-AuNPs exhibited enhanced anti-inflammatory effects on NF-κB-mediated inflammatory network, as well as cell adhesion. Collectively, green synthesis of CGA-AuNPs using bioactive reductants and mechanistic studies based on mass spectrometry may open up new directions in nanomedicine and CGA-AuNPs can be an anti-inflammatory nanomedicine for future applications. FROM THE CLINICAL EDITOR Gold nanoparticles (Au NPs) have been shown to be very useful in many applications due to their easy functionalization capability. In this article, the authors demonstrated a novel method for the synthesis of gold nanoparticles using chlorogenic acid (CGA) as reductants. In-vitro experiments also confirmed biological activity of the resultant gold nanoparticles. Further in-vivo studies are awaited.

Spermidine Protects against Oxidative Stress in Inflammation Models Using Macrophages and Zebrafish

Spermidine is a naturally occurring polyamine compound that has recently emerged with anti-aging properties and suppresses inflammation and oxidation. However, its mechanisms of action on anti-inflammatory and antioxidant effects have not been fully elucidated. In this study, the potential of spermidine for reducing pro-inflammatory and oxidative effects in lipopolysaccharide (LPS)-stimulated macrophages and zebrafish was explored. Our data indicate that spermidine significantly inhibited the production of pro-inflammatory mediators such as nitric oxide (NO) and prostaglandin E2 (PGE2), and cytokines including tumor necrosis factor-α and interleukin-1β in RAW 264.7 macrophages without any significant cytotoxicity. The protective effects of spermidine accompanied by a marked suppression in their regulatory gene expression at the transcription levels. Spermidine also attenuated the nuclear translocation of NF-κB p65 subunit and reduced LPS-induced intracellular accumulation of reactive oxygen species (ROS) in RAW 264.7 macrophages. Moreover, spermidine prevented the LPS-induced NO production and ROS accumulation in zebrafish larvae and was found to be associated with a diminished recruitment of neutrophils and macrophages. Although more work is needed to fully understand the critical role of spermidine on the inhibition of inflammation-associated migration of immune cells, our findings clearly demonstrate that spermidine may be a potential therapeutic intervention for the treatment of inflammatory and oxidative disorders.

Phenyl-β-d-Glucopyranoside Exhibits Anti-inflammatory Activity in Lipopolysaccharide-Activated RAW 264.7 Cells

Phenyl-β-d-glucopyranoside is a component of Phellodendron amurense with anti-cancer and anti-inflammatory activities. In the present study, we investigated the role of phenyl-β-d-glucopyranoside in inflammation using lipopolysaccharide (LPS)-stimulated murine Raw 264.7 macrophages. Phenyl-β-d-glucopyranoside not only inhibited nitric oxide (NO) production but also significantly inhibited the expression of inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2) without inducing cytotoxicity. Phenyl-β-d-glucopyranoside also attenuated proinflammatory cytokines, including tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and other inflammation-related genes, such as IL-6 in a concentration-dependent manner. Furthermore, phenyl-β-d-glucopyranoside abolished increased adhesion, ninjurin 1 (Ninj1) expression, and matrix metalloproteinase (MMP) activity induced by endotoxin treatment. Finally, phenyl-β-d-glucopyranoside inhibited the nuclear translocation of nuclear factor-κB (NF-κB), which is one of the most important transcription factors involved in the inflammatory process. Taken together, phenyl-β-d-glucopyranoside may be beneficial for the prevention and treatment of anti-inflammatory diseases.

Fucoidan inhibits lipopolysaccharide-induced inflammatory responses in RAW 264.7 macrophages and zebrafish larvae

Fucoidan, a sulfated polysaccharide, is an active component found in various species of seaweed. Although this compound has a strong anti-inflammatory activity, the underlying mechanisms exerted by fucoidan have not been fully elucidated. In the present study, the anti-inflammatory effects of fucoidan on lipopolysaccharide (LPS)-stimulated macrophages and zebrafish larvae were examined. The present data indicated that fucoidan significantly suppressed the secretion of pro-inflammatory mediators including nitric oxide (NO ) and prostaglandin E2 (PGE2), and cytokines, such as tumor necrosis factor-α and interleukin-1β in RAW 264.7 macrophages without any significant cytotoxicity, the protective effects of which were accompanied by a marked reduction in their regulatory gene expression at the transcription levels. Fucoidan also inhibited translocation of the nuclear factor-kappa B from the cytoplasm to the nucleus and attenuated LPS-induced production of intracellular reactive oxygen species (ROS) in RAW 264.7 macrophages. Moreover, fucoidan reduced NO and PGE2 production and ROS accumulation in LPS-stimulated zebrafish larvae, which was associated with a diminished recruitment of neutrophils and macrophages. Based on the results of this study, we suggest that fucoidan has excellent potential as a therapeutic agent for inflammatory disorders.

An aqueous extract of Nomura’s jellyfish ameliorates inflammatory responses in lipopolysaccharide-stimulated RAW264.7 cells and a zebrafish model of inflammation

The recent mass emergence of Nomuras jellyfish (Nemopilema nomurai) has caused much economic and environmental damage. However, there is no innovative strategy to dispose of or utilize these jellyfish. Some reports suggest that the jellyfish may be bioactive resources and a source of important compounds with antibacterial activity. Here, we examined the effect of an aqueous extract of Nomuras jellyfish (AENJ) on lipopolysaccharide (LPS)-stimulated Raw 264.7 macrophages and a zebrafish model of inflammation and analyzed the underlying molecular mechanisms. AENJ downregulated inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) mRNA levels in LPS-stimulated Raw 264.7 macrophages, with no apparent cytotoxic effects. However, AENJ had no effect on expression of other inflammation-related genes such as interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and MCP-1. Furthermore, AENJ reduced expression of nerve injury-induced protein 1 (Ninj1), which is an important adhesion molecule, thereby reducing cell adhesion to the extracellular matrix (ECM) in vitro. The inhibitory effect of AENJ on leukocytes was confirmed in LPS-microinjected zebrafish larvae; AENJ reduced the number of the infiltrate accumulating at the site of inflammation. In addition, AENJ suppressed the expression of matrix metalloproteinase-2 (MMP-2) and MMP-9 in LPS-stimulated Raw 264.7 cells. Finally, AENJ blocked nuclear translocation of nuclear factor kappa B (NF-κB), a key transcription factor for inflammatory responses, in Raw 264.7 cells in a dose-dependent manner. Collectively, the data suggest that AENJ inhibits expression of COX and iNOS by blocking NF-κB signaling pathways and suppresses the activity of macrophages by downregulating Ninj1 and MMPs. Therefore, AENJ may be a useful preventive neutraceutical, or therapeutic agent against inflammatory disorders.

Smoking-associated lung cancer prevention by blockade of the beta-adrenergic receptor-mediated insulin-like growth factor receptor activation

Activation of receptor tyrosine kinases (RTKs) is associated with carcinogenesis, but its contribution to smoking-associated lung carcinogenesis is poorly understood. Here we show that a tobacco-specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)-induced insulin-like growth factor 1 receptor (IGF-1R) activation via β-adrenergic receptor (β-AR) is crucial for smoking-associated lung carcinogenesis. Treatment with NNK stimulated the IGF-1R signaling pathway in a time- and dose-dependent manner, which was suppressed by pharmacological or genomic blockade of β-AR and the downstream signaling including a Gβγ subunit of β-AR and phospholipase C (PLC). Consistently, β-AR agonists led to increased IGF-1R phosphorylation. The increase in IGF2 transcription via β-AR, signal transducer and activator of transcription 3 (STAT3), and nuclear factor-kappa B (NF-κB) was associated with NNK-induced IGF-1R activation. Finally, treatment with β-AR antagonists suppressed the acquisition of transformed phenotypes in lung epithelial cells and lung tumor formation in mice. These results suggest that blocking β-AR-mediated IGF-1R activation can be an effective strategy for lung cancer prevention in smokers.

Upcycling of jellyfish (Nemopilema nomurai) sea wastes as highly valuable reducing agents for green synthesis of gold nanoparticles and their antitumor and anti-inflammatory activity

Abstract Due to its tentacle poison and huge body, giant jellyfish (Nemopilema nomurai) poses challenging issues to the environment and ecosystems. Here we developed, upcycling a giant jellyfish extract as a reducing agent, a green synthetic method of gold nanoparticles (JF-AuNPs) which possess biological activities. The colloidal solutions of JF-AuNPs were blue, violet, purple and pink depending on the extract concentration. UV-visible spectra exhibited two surface plasmon resonance bands at 5 4 0 ∼ 550 nm and 810 nm. Spherical shapes with an average size of 35.2 ± 8.7 nm and triangular nanoplates with an average height of 70.5 ± 30.3 nm were observed. A face-centered cubic structure was confirmed by high-resolution X-ray diffraction. JF-AuNPs exhibited significant cytotoxic effect against HeLa cancer cells but not against normal cells such as NIH-3T3 and Raw 264.7 cells. In HeLa cells, JF-AuNPs decreased the phosphorylation of AKT and ERK, which are crucial for cell proliferation. Also, JF-AuNPs decreased NO secretion and iNOS expression levels, resulting in anti-inflammatory effects in LPS-inflamed macrophages. Collectively, we established a green synthesis of anti-tumorigenic and anti-inflammatory JF-AuNPs using the extract of jellyfish sea wastes. Thus, beneficial effects of JF-AgNPs must be weighed in further studies in vivo and it can be potent nanomedicine for future applications.

A synthetic Nitraria alkaloid, isonitramine protects pancreatic β-cell and attenuates postprandial hyperglycemia

OBJECTIVE The extracts of Nitraria genus are composed of Nitraria alkaloids and have been used traditionally as a hypoglycemic medicine. However, the efficacy and precise mechanism of Nitraria alkaloids remain largely unknown. METHODS Previously, we reported the total synthesis of (+)-isonitramine, one of Nitraria alkaloids. In this study, we investigated the anti-diabetic potential of isonitramine in diabetes mellitus and its underlying molecular mechanism in carbohydrate catabolism in vitro and in vivo. RESULTS Isonitramine exerted significant inhibitory effect on α-glucosidases but not α-amylase in vitro. In zebrafish, isonitramine alleviated the streptozotocin (STZ)-induced postprandial hyperglycemia and protected the pancreatic damages against alloxan-induced oxidative stress in vivo. Also, isonitramine induced insulin without any toxicities and downregulated phosphoenolpyruvate carboxykinase (PEPCK), which catalyzes the first committed step in gluconeogenesis. CONCLUSION Taken together, isonitramine inhibited α-glucosidase activity and PEPCK expression, while increased insulin expression, resulting in attenuating the postprandial hyperglycemia. Also, isonitramine protected the pancreas from ROS-mediated toxicities. Therefore, isonitramine may be a new drug candidate for the treatment of diabetes mellitus.