Original article Effects of hypothermia on ex vivo microglial production of pro- and anti-inflammatory cytokines and nitric oxide in hypoxic-ischemic brain-injured mice

Abstract Background Macrophages play a crucial role in inflammation. Astilbe chinensis is one of perennial herbs belonging to the genus Astilbe. Plants in the genus have been used for pain, headaches, arthralgia, and chronic bronchitis. However, the effect of A.chinensis on inflammation remains unclear. To study the anti-inflammatory action of A.chinensis ethanol extract (ACE), we investigated the effect of ACE on the production of pro-inflammatory mediators and cytokines in macrophages. Methods We evaluated the effectiveness of ACE in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages and thioglycollate (TG)-elicited peritoneal macrophages from male C57BL/6 mice. We measured the levels of pro-inflammatory mediators and cytokines, and examined the anti-inflammatory actions of ACE on nuclear factor κB (NF-κB) pathway in the macrophages. Western blot analysis and immunofluorescence microscopy were used to determine protein level and translocation, respectively. Results ACE suppressed the output of nitric oxide (NO), prostaglandin E2 (PGE2), and pro-inflammatory cytokines in stimulated macrophages via inhibiting the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) proteins. ACE suppressed mRNA expression of pro-inflammatory cytokines such as interleukin (IL)-6 and tumor necrosis factor-alpha (TNF-α). We examined the efficacies of ACE on NF-κB activation by measuring the expressions including IκB kinase (IKK), inhibitor of κB (IκB), and nuclear p65 proteins. In addition, the inhibition of NF-κB p65’s translocation was determined with immunofluorescence assay. Conclusion Our findings manifested that ACE inhibited LPS or TG-induced inflammation by blocking the NF-κB signaling pathway in macrophages. It indicated that ACE is a potential therapeutic mean for inflammation and related diseases.

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Hookworm-Derived Metabolites Suppress Pathology in a Mouse Model of Colitis and Inhibit Secretion of Key Inflammatory Cytokines in Primary Human Leukocytes

Infection and Immunity ◽

10.1128/iai.00851-18 ◽

2019 ◽

Vol 87 (4) ◽

Cited By ~ 10

Author(s):

Phurpa Wangchuk ◽

Catherine Shepherd ◽

Constantin Constantinoiu ◽

Rachael Y. M. Ryan ◽

Konstantinos A. Kouremenos ◽

...

Keyword(s):

Fatty Acids ◽

Inflammatory Cytokines ◽

Mononuclear Cells ◽

Ex Vivo ◽

Clinical Signs ◽

Gas Chromatography Mass Spectrometry ◽

Trinitrobenzenesulfonic Acid ◽

Human Leukocytes ◽

Polar Metabolites ◽

Anti Inflammatory

ABSTRACT Iatrogenic hookworm therapy shows promise for treating disorders that result from a dysregulated immune system, including inflammatory bowel disease (IBD). Using a murine model of trinitrobenzenesulfonic acid-induced colitis and human peripheral blood mononuclear cells, we demonstrated that low-molecular-weight metabolites derived from both somatic extracts (LMWM-SE) and excretory-secretory products (LMWM-ESP) of the hookworm, Ancylostoma caninum, display anti-inflammatory properties. Administration to mice of LMWM-ESP as well as sequentially extracted fractions of LMWM-SE using both methanol (SE-MeOH) and hexane-dichloromethane-acetonitrile (SE-HDA) resulted in significant protection against T cell-mediated immunopathology, clinical signs of colitis, and impaired histological colon architecture. To assess bioactivity in human cells, we stimulated primary human leukocytes with lipopolysaccharide in the presence of hookworm extracts and showed that SE-HDA suppressed ex vivo production of inflammatory cytokines. Gas chromatography-mass spectrometry (MS) and liquid chromatography-MS analyses revealed the presence of 46 polar metabolites, 22 fatty acids, and five short-chain fatty acids (SCFAs) in the LMWM-SE fraction and 29 polar metabolites, 13 fatty acids, and six SCFAs in the LMWM-ESP fraction. Several of these small metabolites, notably the SCFAs, have been previously reported to have anti-inflammatory properties in various disease settings, including IBD. This is the first report showing that hookworms secrete small molecules with both ex vivo and in vivo anti-inflammatory bioactivity, and this warrants further exploration as a novel approach to the development of anti-inflammatory drugs inspired by coevolution of gut-dwelling hookworms with their vertebrate hosts.

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Evaluation of the Anti-inflammatory and Anti-tumor Effect of Ipomoea obscura (L) and Its Mode of Action Through the Inhibition of Pro Inflammatory Cytokines, Nitric Oxide and COX-2

Inflammation ◽

10.1007/s10753-010-9221-4 ◽

2010 ◽

Vol 34 (3) ◽

pp. 171-183 ◽

Cited By ~ 18

Author(s):

T. P. Hamsa ◽

Girija Kuttan

Keyword(s):

Nitric Oxide ◽

Inflammatory Cytokines ◽

Mode Of Action ◽

Cox 2 ◽

Anti Inflammatory ◽

Pro Inflammatory Cytokines ◽

Ipomoea Obscura

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Anti-Inflammatory Activity of Sonchus oleraceus Extract in Lipopoly saccharide-Stimulated RAW264.7 Cells

Biomedical & Pharmacology Journal ◽

10.13005/bpj/1546 ◽

2018 ◽

Vol 11 (4) ◽

pp. 1755-1761

Author(s):

Eun-Jin Yang ◽

Sungchan Jang ◽

Kwang Hee Hyun ◽

Eun-Young Jung ◽

Seung-Young Kim ◽

...

Keyword(s):

Nitric Oxide ◽

Inflammatory Cytokines ◽

Inflammatory Activity ◽

Raw264.7 Cells ◽

Supporting Evidence ◽

Dependent Manner ◽

Sonchus Oleraceus ◽

Raw264.7 Macrophages ◽

Anti Inflammatory ◽

Pro Inflammatory Cytokines

The anti-inflammatory activity and non-toxicity of Sonchus oleraceus extract (J6) were tested by measuring its effect on the levels of nitric oxide (NO), prostaglandin E2 (PGE2), and the pro-inflammatory cytokines, interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α), in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages. We treated the RAW264.7 cells with various concentrations (50, 100, or 200 μg/mL) of J6. Our results showed that J6 inhibited the production of NO, PGE2, and pro-inflammatory cytokines in a concentration-dependent manner, without compromising cell viability. In addition, we provided supporting evidence that the inhibitory activity of J6 on the production of NO and PGE2 occurred via the downregulation of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), respectively. Our findings suggest that J6 is a new source for anti-inflammatory drugs and ingredients for healthcare products that include functional cosmetics.

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Anti‐inflammatory effects of Laminaria japonica in association with the inhibition of nitric oxide and pro‐inflammatory cytokines production

The FASEB Journal ◽

10.1096/fasebj.26.1_supplement.lb547 ◽

2012 ◽

Vol 26 (S1) ◽

Author(s):

Young Woo Kim ◽

Je Cho ◽

Sang Chan Kim

Keyword(s):

Nitric Oxide ◽

Inflammatory Cytokines ◽

Laminaria Japonica ◽

Anti Inflammatory ◽

Pro Inflammatory Cytokines

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Polyene Phosphatidylcholine Interacting with TLR-2 Prevents the Synovial Inflammation via Inactivation of MAPK and NF-κB Pathways

10.21203/rs.3.rs-1078229/v1 ◽

2021 ◽

Author(s):

Zixuan Xu ◽

Wenting Hao ◽

Daxiang Xu ◽

Yan He ◽

Ziyi Yan ◽

...

Keyword(s):

Inflammatory Cytokines ◽

Ex Vivo ◽

Mapk Pathway ◽

Synovial Fibroblasts ◽

Underlying Mechanism ◽

Inhibitory Effect ◽

Synovial Inflammation ◽

Joint Disease ◽

Drug Candidate ◽

Anti Inflammatory

Abstract Rheumatoid arthritis (RA) is a chronic autoimmune joint disease that causes cartilage and bone damage or even disability, seriously endangering human health. Chronic synovial inflammation has been shown to play a vital role in the disease sustainability. Therefore, down-regulation of synovial inflammation is considered to be an effective discipline for RA therapy. Polyene phosphatidylcholine (PPC) is a hepatoprotective agent, which was observed to inhibit inflammation in macrophages and prevent collagen-induced arthritis (CIA) of rats in our previous study. However, the underlying mechanism remains unclear. The present study further reported that PPC can inhibit the synovial inflammation. In lipopolysaccharide (LPS)-stimulated primary synovial fibroblasts (SFs) of mice, PPC significantly decreased pro-inflammatory cytokines production while increasing anti-inflammatory cytokines level. In this process, PPC down-regulated the expression of TLR-2 and their downstream signaling molecules such as MyD88, p-ERK1/2, p-JNK1/2, p-P38 in the MAPK pathway and p-IκBα and NF-κB-p65 in NF-kB pathway. Moreover, the inhibitory effect of PPC on the above molecules and cytokines was weakened after the use of TLR-2 agonist Pam3CSK4. However, PPC lost its anti-inflammatory effect and showed an activation of MAPK and NF-kB pathways in the TLR-2-/- primary SFs after exposure to LPS. Furthermore, these results were confirmed in the SFs from the CIA mouse ex vivo. Collectively, this study demonstrated that PPC can alleviate synovial inflammation through TLR-2 mediated MAPK and NF-κB pathways, which can be proposed to be a potential drug candidate for RA therapy.

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N-Butanol Extract of Gastrodia elata Suppresses Inflammatory Responses in Lipopolysaccharide-Stimulated Macrophages and Complete Freund’s Adjuvant- (CFA-) Induced Arthritis Rats via Inhibition of MAPK Signaling Pathway

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2020/1658618 ◽

2020 ◽

Vol 2020 ◽

pp. 1-11

Author(s):

Peng He ◽

Yiwen Hu ◽

Changzhao Huang ◽

Xi Wang ◽

Heng Zhang ◽

...

Keyword(s):

Nitric Oxide ◽

Therapeutic Effect ◽

Inflammatory Cytokines ◽

Gastrodia Elata ◽

Butanol Extract ◽

Freund’S Adjuvant ◽

Cox 2 ◽

Freund's Adjuvant ◽

Anti Inflammatory ◽

Raw264.7 Macrophage

Gastrodia elata is a traditional herbal medicine that has been used for centuries to treat rheumatism. Previous studies have confirmed that ethanol extracts of Gastrodia elata have anti-inflammatory and antioxidant activities, and the n-butanol fraction exerts a higher inhibitory effect. However, the in vivo anti-inflammatory effects of Gastrodia elata have not been evaluated. Thus, we assessed the therapeutic effect of the n-butanol extract of Gastrodia elata (BGE) on complete Freund’s adjuvant- (CFA-) induced arthritis rats which were separated into six groups (NOR; MODEL; CFA + dexamethasone (DEX); CFA + 25, 50, 100 mg/kg BGE). The paw swelling, joint radiology, and histology were used to analyze the effect of BGE on delaying the progression of rheumatoid arthritis. Furthermore, serum levels of inflammatory cytokines were analyzed via ELISA. In addition, the effect of BGE on nitric oxide (NO) production, expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2(COX-2), and inflammatory cytokines were detected in lipopolysaccharide- (LPS-) stimulated RAW264.7 macrophage cells. Lastly, the impacts of BGE on the activation of the mitogen-activated protein kinases (MAPK) pathway in CFA rats and LPS-stimulated RAW264.7 macrophage were examined by western blot analysis. The results show that BGE can significantly reduce paw swelling without losing the body weight of rats. Imaging assessment confirms that BGE can protect cartilage from destruction, as well as reducing inflammatory cell infiltration and synovial proliferation. Moreover, BGE suppresses the production of inflammatory cytokines in serum and inhibits the activation of the phosphorylation of p38 and ERK in CFA rats. BGE was also demonstrated to decrease the production of NO and inflammatory cytokines in LPS-stimulated RAW264.7 cells. The effect of BGE in LPS-induced expression leads to reduced p38 and ERK phosphorylation and also downregulates the protein expression of iNOS and COX-2. Taken together, BGE exhibits a potential therapeutic effect on CFA rats, and its anti-inflammatory and antioxidant effects were possibly exerted by regulation of ERK/p38MAPK.

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Ex Vivo and In Vivo Evidence of Anti-Inflammatory Activity of P-aminophenol and Salicylate Derivatives

Current Bioactive Compounds ◽

10.2174/1573407215666190211150700 ◽

2020 ◽

Vol 16 (5) ◽

pp. 593-605

Author(s):

André F. Vilvert ◽

Marcus Vinícius P.S. Nascimento ◽

Rosivaldo dos S. Borges ◽

Eduardo M. Dalmarco

Keyword(s):

Nitric Oxide ◽

Interleukin 6 ◽

Ex Vivo ◽

Inflammatory Activity ◽

Potential Candidate ◽

Antiinflammatory Drugs ◽

Gentisic Acid ◽

Anti Inflammatory ◽

Inflammatory Effect

Background: Paracetamol (p-aminophenol) and salicylates are nonsteroidal antiinflammatory drugs that are widely used in the general population. The adverse effects of both drugs continue to be a focus of the pharmaceutical industry in the development of new molecules that will increase treatment safety. In this context, we tested nine compounds derived from paracetamol and salicylates, synthesized in our laboratory, for their safety and ex vivo and in vivo anti-inflammatory activity. Methods: We analyzed the cytotoxicity of the compounds in ex vivo mice neutrophils, and their ability to inhibit the production of pro-inflammatory mediators (nitric oxide and interleukin-6) after stimulating with LPS. Next, in the selected molecules, we evaluated the anti-inflammatory effect on an in vivo inflammatory model of acute lung injury in mice. All nine compounds were also submitted to the cytotoxicity assay, like the original compounds. Results: None of the compounds showed cytotoxicity under the cells used. However, of the initial compounds, only five demonstrated anti-inflammatory effect, inhibiting Nitric Oxide (NO) and interleukin 6 (IL-6) production by neutrophils stimulated with Lipopolysaccharide (LPS). After this initial trial, four modified compounds were able to reduce leukocyte migration and fluid leakage in the bronchoalveolar lavage of mice. However, only the compound 5a1, derived from the esterification of gentisic acid, was able to significantly inhibit the levels of all pro-inflammatory cytokines and increase the levels of antiinflammatory cytokines evaluated. Conclusion: In conclusion, all compounds showed a good safety profile, and many of them had an antiinflammatory effect. However, the compound derived from gentisic acid is highlighted for its significant effects ex vivo and in vivo and in this context, we believe that this compound is a potential candidate for the development of a new anti-inflammatory drug.

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Anti-Inflammatory Effect of Pineapple Rhizome Bromelain through Downregulation of the NF-κB- and MAPKs-Signaling Pathways in Lipopolysaccharide (LPS)-Stimulated RAW264.7 Cells

Current Issues in Molecular Biology ◽

10.3390/cimb43010008 ◽

2021 ◽

Vol 43 (1) ◽

pp. 93-106

Author(s):

Orapin Insuan ◽

Phornphimon Janchai ◽

Benchaluk Thongchuai ◽

Rujirek Chaiwongsa ◽

Supaporn Khamchun ◽

...

Keyword(s):

Nitric Oxide ◽

Signaling Pathways ◽

Inflammatory Cytokines ◽

Molecular Mechanisms ◽

Raw264.7 Cells ◽

Nuclear Factor ◽

Amino Terminal ◽

Cox 2 ◽

Anti Inflammatory ◽

Pro Inflammatory Cytokines

Bromelain is a mixture of proteolytic enzymes derived from pineapple (Ananas comosus) fruit and stem possessing several beneficial properties, particularly anti-inflammatory activity. However, the molecular mechanisms underlying the anti-inflammatory effects of bromelain are unclear. This study investigated the anti-inflammatory effects and inhibitory molecular mechanisms of crude and purified rhizome bromelains on lipopolysaccharide (LPS)-induced inflammation in RAW 264.7 macrophage cells. RAW264.7 cells were pre-treated with various concentrations of crude bromelain (CB) or purified bromelain (PB), and then treated with LPS. The production levels of pro-inflammatory cytokines and mediators, including nitric oxide (NO), interleukin (IL)-6, and tumor necrosis factor (TNF)-α were determined by Griess and ELISA assays. The expressions of inducible nitric oxide synthetase (iNOS), cyclooxygenase (COX)-2, nuclear factor kappa B (NF-κB), and mitogen-activated protein kinases (MAPKs)-signaling pathway-related proteins were examined by western blot analysis. The pre-treatment of bromelain dose-dependently reduced LPS-induced pro-inflammatory cytokines and mediators, which correlated with downregulation of iNOS and COX-2 expressions. The inhibitory potency of PB was stronger than that of CB. PB also suppressed phosphorylated NF-κB (p65), nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor alpha, extracellular signal-regulated kinases, c-Jun amino-terminal kinases, and p38 proteins in LPS-treated cells. PB then exhibited potent anti-inflammatory effects on LPS-induced inflammatory responses in RAW264.7 cells by inhibiting the NF-κB and MAPKs-signaling pathways.

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