Optimization of an anti-inflammatory screening model on the RAW 264.7 macrophage cell

Inflammation is the response of living tissues to the injury. Prolonged or inappropriate inflammation has been involved with many diseases such as, cancer, diabetes, heart disease… These days, inflammation has been treated by nonsteroidal and steroidal anti-inflammatory drugs, which have a lot of side effects. It opens the need and interest of new drugs discovery. Particularly, scientific and pharmaceutical communities show great interest in finding new anti-inflammatory compounds in traditional medicinal plants. This study aimed to optimize an in vitro anti-inflammatory model. Cell heterogeneity, cell density, LPS concentration and LPS incubation time were chosen to optimize the production of nitric oxide (NO) in RAW 264.7 macrophage cells. Our results show that 104 cells/well, FBS 1 %, starvation for 6 h, LPS 0.5 µg/mL in 24 h were optimized parameters in the model. Then, extracts from Hedyotis capitellata, a traditional medicinal plant used by K’ho minority, Bidoup Nuiba national park, Lam Dong province, Vietnam, was chosen to evaluate the in vitro antiinflammatory model. The anti-inflammatory activity was tested by measuring the production of NO in lipopolysaccharide-activated RAW 264.7 macrophage cells. The experimental data indicate that the extracts of this plant decreased NO production in LPS-stimulated RAW 264.7, particularly, the petroleum ether fraction at the concentration of 23.8 µg/mL inhibited NO production by 128.20 %; whereas dexamethasone 50 µM inhibited NO accumulation to 112 %.

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Identification and Characterization of Edible Cricket Peptides on Hypertensive and Glycemic In Vitro Inhibition and Their Anti-Inflammatory Activity on RAW 264.7 Macrophage Cells

Nutrients ◽

10.3390/nu12113588 ◽

2020 ◽

Vol 12 (11) ◽

pp. 3588

Author(s):

Felicia Hall ◽

Lavanya Reddivari ◽

Andrea M. Liceaga

Keyword(s):

De Novo ◽

Binding Energies ◽

Raw 264.7 ◽

Sequential Fractionation ◽

Enzyme Active Site ◽

Macrophage Cells ◽

Raw 264.7 Macrophage ◽

Raw 264.7 Macrophage Cells ◽

Anti Inflammatory

Recent studies continue to demonstrate the potential of edible insects as a protein base to obtain bioactive peptides applicable for functional food development. This study aimed at identifying antihypertensive, anti-glycemic, and anti-inflammatory peptides derived from the in vitro gastrointestinal digests of cricket protein hydrolysates. After sequential fractionation, the protein digest subfraction containing the lowest molecular weight (<0.5 kDa), hydrophobic (C18) and cationic peptides (IEX) was found responsible for the most bioactivity. The cationic peptide fraction significantly reduced (p < 0.05) α-amylase, α-glucosidase, and angiotensin converting enzyme (ACE) activity in vitro, and also inhibited the expression of NF-κB in RAW 264.7 macrophage cells. A total of 28 peptides were identified with mass spectrometry (LC–MS/MS) and de novo sequencing from the potent fraction. Three novel peptides YKPRP, PHGAP, and VGPPQ were chosen for the molecular docking studies. PHGAP and VGPPQ exhibited a higher degree of non-covalent interactions with the enzyme active site residues and binding energies comparable to captopril. Results from this study demonstrate the bioactive potential of edible cricket peptides, especially as ACE inhibitors.

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Anti-inflammatory and anti-allergic activities of Skipjack tuna (Katsuwonus pelamis) dark muscle hydrolysates evaluated in cell culture model

Functional Foods in Health and Disease ◽

10.31989/ffhd.v9i7.618 ◽

2019 ◽

Vol 9 (7) ◽

pp. 446

Author(s):

Worrapanit Chansuwan ◽

Chutha Takahashi Upunqui ◽

Pavinee Chinachoti

Keyword(s):

Molecular Weight ◽

No Production ◽

Raw 264.7 ◽

Skipjack Tuna ◽

Dark Muscle ◽

Macrophage Cells ◽

Raw 264.7 Macrophage ◽

Raw 264.7 Macrophage Cells ◽

Anti Inflammatory ◽

Anti Inflammation

Background: Oxidative stress and inflammation are inextricably linked and play major roles in the onset and development of Non-communicable diseases (NCD) which are the most common cause of death and disability in modern world. Hydrolyzed proteins have also been suggested to be used to manage adverse food allergic reaction. Therefore, this study aimed to investigate anti-inflammatory and anti-allergy activities of dark muscle tuna hydrolysates using biological cell line systems as a function of enzyme, the extent of hydrolysis and molecular weight range.Methods: Dark muscle tuna hydrolysates were prepared with two different enzyme types; Alcalase and Flavourzyme. Anti-inflammation activity was measured by inhibitory effect of nitric oxide (NO) production on lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophage cells. Anti-allergy was determined from ability of hydrolysates to inhibit b-hexsosaminidase (b-HEX) release from RBL-2H3 mast cells. Cytotoxicity was also investigated in both RAW 264.7 macrophage cells and RBL-2H3 mast cells.Results: No cytotoxic effect on RAW 264.7 macrophage cells and RBL-2H3 mast cells was observed. The NO inhibition and b-HEX release were found significant in dose dependent manner (p<0.05). Alcalase hydrolysates demonstrated greater anti-inflammatory and anti-allergic activities than Flavourzyme hydrolysates (p<0.05). IC50 of both effects were lower than the unhydrolyzed control, > 45.44 mg/ml for NO inhibition and > 65.23 mg/ml for b-HEX release inhibition. These effects increased with the extent of hydrolysis and enzyme concentration. The peptide of lowest molecular weight range (< 3 KDa) was highest in anti-inflammatory and anti-allergic actions. Reducing secretion of TNF-a, IL-6 and IL-1b was found greater in Alcalase hydrolysate than Flavourzyme one.Conclusions: Skipjack tuna dark muscle hydrolysates from Alcalase resulted in peptides with anti-inflammation activity, as determined by NO production in LPS-stimulated RAW 264.7 macrophage cells and anti-allergic properties as measured by a suppression of degranulation of sensitized RBL-2H3 cells. Anti-inflammatory effect may be due to their anti-oxidative capacity and relevant inflammatory factors attenuated with hydrolysate by reducing secretion of pro-inflammatory cytokine (TNF-a, IL-6 and IL-1b). Inhibition of b-HEX release by peptides may be due to membrane-stabilizing action or/and blockade of IgE antibody at fragment region.Keywords: Skipjack tuna, anti-inflammation, enzymatic hydrolysate, dark muscle, anti-allergy

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Anti-inflammatory Effects of Madecassic Acid via the Suppression of NF-κB Pathway in LPS-Induced RAW 264.7 Macrophage Cells

Planta Medica ◽

10.1055/s-0029-1186142 ◽

2009 ◽

Vol 76 (03) ◽

pp. 251-257 ◽

Cited By ~ 53

Author(s):

Jong-Heon Won ◽

Ji-Sun Shin ◽

Hee-Juhn Park ◽

Hyun-Ju Jung ◽

Duck-Jae Koh ◽

...

Keyword(s):

Raw 264.7 ◽

Macrophage Cells ◽

Raw 264.7 Macrophage ◽

Raw 264.7 Macrophage Cells ◽

Anti Inflammatory

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Antioxidant and Anti-Inflammatory Effects of Defatted Rice Bran (Oryza SativaL.) Protein Hydrolysates on Raw 264.7 Macrophage Cells

Journal of Food Biochemistry ◽

10.1111/jfbc.12266 ◽

2016 ◽

Vol 40 (6) ◽

pp. 731-740 ◽

Cited By ~ 17

Author(s):

Tanatorn Saisavoey ◽

Papassara Sangtanoo ◽

Onrapak Reamtong ◽

Aphichart Karnchanatat

Keyword(s):

Rice Bran ◽

Protein Hydrolysates ◽

Raw 264.7 ◽

Defatted Rice Bran ◽

Macrophage Cells ◽

Raw 264.7 Macrophage ◽

Raw 264.7 Macrophage Cells ◽

Anti Inflammatory

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Haloperidol and Risperidone at high concentrations activate an in vitro inflammatory response of RAW 264.7 macrophage cells by induction of apoptosis and modification of cytokine levels

Psychopharmacology ◽

10.1007/s00213-015-4079-7 ◽

2015 ◽

Vol 233 (9) ◽

pp. 1715-1723 ◽

Cited By ~ 18

Author(s):

Ivo Emílio da Cruz Jung ◽

Alencar Kolinski Machado ◽

Ivana Beatrice Mânica da Cruz ◽

Fernanda Barbisan ◽

Verônica Farina Azzolin ◽

...

Keyword(s):

Inflammatory Response ◽

Raw 264.7 ◽

Macrophage Cells ◽

Raw 264.7 Macrophage ◽

Cytokine Levels ◽

Raw 264.7 Macrophage Cells ◽

Induction Of Apoptosis ◽

High Concentrations

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Anti-Inflammatory Effects of Formononetin 7-O-phosphate, a Novel Biorenovation Product, on LPS-Stimulated RAW 264.7 Macrophage Cells

Molecules ◽

10.3390/molecules24213910 ◽

2019 ◽

Vol 24 (21) ◽

pp. 3910 ◽

Cited By ~ 2

Author(s):

Min-Seon Kim ◽

Jin-Soo Park ◽

You Chul Chung ◽

Sungchan Jang ◽

Chang-Gu Hyun ◽

...

Keyword(s):

Biological Properties ◽

In Vitro Assays ◽

Raw 264.7 Cells ◽

No Production ◽

Raw 264.7 ◽

Dependent Manner ◽

Macrophage Cells ◽

Anti Inflammatory ◽

Reduced Toxicity

Biorenovation is a microbial enzyme-catalyzed structural modification of organic compounds with the potential benefits of reduced toxicity and improved biological properties relative to their precursor compounds. In this study, we synthesized a novel compound verified as formononetin 7-O-phosphate (FMP) from formononetin (FM) using microbial biotransformation. We further compared the anti-inflammatory properties of FMP to FM in lipopolysaccharide (LPS)-treated RAW264.7 macrophage cells. We observed that cell viabilities and inhibitory effects on LPS-induced nitric oxide (NO) production were greater in FMP-treated RAW 264.7 cells than in their FM-treated counterparts. In addition, FMP treatment suppressed the production of proinflammatory cytokines such as prostaglandin-E2 (PGE2), interleukin-6 (IL-6), and interleukin-1β (IL-1β) in a dose-dependent manner and concomitantly decreased the mRNA expression of inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2). We also found that FMP exerted its anti-inflammatory effects through the downregulation of the extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and nuclear factor kappa B (NF-κB) signaling pathways. In conclusion, we generated a novel anti-inflammatory compound using biorenovation and demonstrated its efficacy in cell-based in vitro assays.

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