Cyclooxygenase-2 mRNA Expression Research Articles

Antioxidant and Anti-Inflammatory Activity on LPS-Stimulated RAW 264.7 Macrophage Cells of White Mulberry (Morus alba L.) Leaf Extracts.

The white mulberry (Morus alba L.) is widely used as a medicinal plant in Asia. In this study, the bioactive compounds of ethanolic extracts of white mulberry leaves from the Sakon Nakhon and Buriram cultivars were evaluated. The ethanolic extracts of mulberry leaves from the Sakon Nakhon cultivar showed the highest total phenolic content of 49.68 mg GAE/g extract and antioxidant activities of 4.38 mg GAE/g extract, 4.53 mg TEAC/g extract, and 92.78 mg FeSO4/g extract using 2,2 diphenyl-1-picrylhydrazyl (DPPH), 2,20-azinobis-(3-ethylbenzothiazolin-6-sulfonic acid) (ABTS), and ferric reducing antioxidant power (FRAP) assays, respectively. The resveratrol and oxyresveratrol compounds in mulberry leaves were also investigated by high-performance liquid chromatography (HPLC). The mulberry leaf extracts from the Sakon Nakhon and Buriram cultivars showed oxyresveratrol contents of 1.20 ± 0.04 mg/g extract and 0.39 ± 0.02 mg/g extract, respectively, whereas resveratrol was not detected. It was also found that the potent anti-inflammatory properties of mulberry leaf extracts and its compounds, resveratrol and oxyresveratrol, suppressed the LPS-stimulated inflammatory responses in RAW 264.7 macrophage cells by significantly reducing nitric oxide production in a concentration-dependent manner. These compounds further inhibited interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) production and suppressed the mRNA and protein expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in LPS-stimulated RAW 264.7 macrophage cells. Therefore, it is established that mulberry leaf extract and its bioactive compounds contribute to its anti-inflammatory activity.

Fermented Kamut Sprout Extract Decreases Cell Cytotoxicity and Increases the Anti-Oxidant and Anti-Inflammation Effect.

Kamut sprouts (KaS) contain several biologically active compounds. In this study, solid-state fermentation using Saccharomyces cerevisiae and Latilactobacillus sakei was used to ferment KaS (fKaS-ex) for 6 days. The fKaS-ex showed a 26.3 mg/g dried weight (dw) and 46.88 mg/g dw of polyphenol and the β-glucan contents, respectively. In the Raw264.7 and HaCaT cell lines, the non-fermented KaS (nfKaS-ex) decreased cell viability from 85.3% to 62.1% at concentrations of 0.63 and 2.5 mg/mL, respectively. Similarly, the fKaS-ex decreased cell viability, but showed more than 100% even at 1.25 and 5.0 mg/mL concentrations, respectively. The anti-inflammatory effect of fKaS-ex also increased. At 600 µg/mL, the fKaS-ex exhibited a significantly higher ability to reduce cytotoxicity by suppressing COX-2 and IL-6 mRNA expressions as well as that for IL-1β mRNA. In summary, fKaS-ex exhibited significantly lower cytotoxicity and increased anti-oxidant and anti-inflammatory properties, indicating that fKaS-ex is beneficial for use in food and other industries.

The pivotal role of Bifida Ferment Lysate on reinforcing the skin barrier function and maintaining homeostasis of skin defenses in vitro.

The semiactive or inactive probiotics or their extracts used in dermatology have interesting properties to ameliorate signs of irritated skin and enhance the skin barrier. Bifidobacterium, as the most common probiotics, which has been found to be effective in reducing acne and improving the skin barrier function of atopic dermatitis. Bifida Ferment Lysate (BFL) can be obtained from Bifidobacterium by fermentation and extraction. In this study, we investigated the effect of a topically used BFL on the skin using in vitro evaluation methods. The results showed that upregulation of skin physical barrier gene (FLG, LOR, IVL, TGM1, and AQP3) and antimicrobial peptide gene (CAMP and hBD-2) in HaCaT cells by BFL might be responsible for skin barrier resistance. In addition, BFL had strong antioxidant properties representing a dose-dependent increasing of the scavenging capacity of DPPH, ABTS, hydroxyl, and superoxide radicals. BFL treatment also fundamentally inhibited the intracellular ROS and MDA production and improved the activities of antioxidant enzymes (CAT and GSH-Px) in H2 O2 -stimulated HaCaT cells. As a good immunomodulatory factor, BFL efficiently decreased the secretion of IL-8 and TNF-α cytokines, and COX-2 mRNA expression in LPS-induced THP-1 macrophages. BFL can strengthen the skin barrier function and stimulate skin barrier resistance, to reinforce the skin against oxidative stress and inflammatory stimuli.

Antioxidative and Anti-Inflammatory Activities of Rosebud Extracts of Newly Crossbred Roses.

Oxidative stress and inflammation are basic pathogenic factors involved in tissue injury and pain, as well as acute and chronic diseases. Since long-term uses of synthetic steroids and non-steroidal anti-inflammatory drugs (NSAIDs) cause severe adverse effects, novel effective materials with minimal side effects are required. In this study, polyphenol content and antioxidative activity of rosebud extracts from 24 newly crossbred Korean roses were analyzed. Among them, Pretty Velvet rosebud extract (PVRE) was found to contain high polyphenols and to show in vitro antioxidative and anti-inflammatory activities. In RAW 264.7 cells stimulated with lipopolysaccharide (LPS), PVRE down-regulated mRNA expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), and thereby decreased nitric oxide (NO) and prostaglandin E2 (PGE2) production. In a subcutaneous air-pouch inflammation model, treatment with PVRE decreased λ-carrageenan-induced tissue exudation, infiltration of inflammatory cells, and inflammatory cytokines such as tumor necrosis factor-α and interleukin-1β concentrations, as achieved with dexamethasone (a representative steroid). Notably, PVRE also inhibited PGE2, similar to dexamethasone and indomethacin (a representative NSAID). The anti-inflammatory effects of PVRE were confirmed by microscopic findings, attenuating tissue erythema, edema, and inflammatory cell infiltration. These results indicate that PVRE exhibits dual (steroid- and NSAID-like) anti-inflammatory activities by blocking both the iNOS-NO and COX-2-PG pathways, and that PVRE could be a potential candidate as an anti-inflammatory material for diverse tissue injuries.

Diosgenin, a steroidal sapogenin, arrests arthritis through modulation of inflammatory cytokines and oxidative stress biomarkers in Wistar rats.

Diosgenin (DGN) is a well-known steroidal sapogenin that is obtained from the hydrolysis of dioscin. The current research aimed to explore the anti-inflammatory and anti-arthritic potential of DGN alone and in combination with methotrexate (MTX). The in-vitro antioxidant, and anti-arthritic potential was assessed by protein denaturation and Human red blood cell membrane stabilization assays. The in-vivo anti-inflammatory effect was examined by carrageenan-induced paw edema and xylene-induced ear edema methods. The arthritis was induced in Wistar rats by inoculation of 0.1ml Complete Freund's adjuvant in the left hind paw at day 1. The arthritic animals received MTX 1mg/kg as standard, DGN at 5, 10, 20mg/kg, and a combination treatment (DGN 20mg/kg + MTX) was administered orally from 8 to 28th day while normal and disease control received normal saline. DGN at 1600μg/ml exhibited the highest in-vitro activities in contrast to other tested concentrations. DGN at 20mg/kg exhibited the maximum (p < 0.05-0.0001) inhibition of inflammation in carrageenan and xyleneinduced edema models. Treatment with DGN and MTX alone and in combination significantly reduced the paw diameter, body weight, arthritic index, and pain. It restored altered blood parameters and oxidative stress biomarkers in contrast to the diseased control rats. DGN profoundly (P < 0.0001) downregulated mRNA expression of TNF-α, IL-1β, NF-ĸβ, and COX-2 while upregulated IL-4 and -10 in treated rats. The combination of DGN with MTX showed the highest therapeutic efficacy than individual therapy, so it can be used as an adjunct for rheumatoid arthritis treatment.

Fucoidan (A Sulfated Polysaccharide) and Cerebroprotein in Combination Alleviate the Neuroinflammation-mediated Neural Damage and Functional Deficits in the Focal Cerebral Ischemia Model of Rat

Cerebral ischemic reperfusion injury could emanate a cascade of events ensuing in neural death and severe neurobehavioural deficits. The currently available interventions have failed to target the multimodal, interlinked mechanisms that operate cerebral ischemia-induced damage and functional loss. So an integrative intervention has become a mandate to overcome the deleterious mechanisms involved in cerebral ischemic pathophysiology. In this study, adult male Sprague dawley rats were exposed to 2 hours of right middle cerebral artery occlusion (rMCAo) followed by reperfusion, and the intervention group received Fucoidan alone at a dose of 50 mg/kg, i.p (intraperitoneal), Cerebrolysin alone at a dose of 2.5 mg/kg body weight and the combination of both. The sham rats were exposed to surgical procedures, except for the rMCAo. The assessments of the groups were made 24 h after the rMCAo. The stand-alone treatment with Fucoidan, Cerebrolysin has shown a better outcome in the neurobehavioral and, histopathological assessments and the combination has made a significant reduction in the neurological deficits and the infarct volume when compared to the standalone groups. The BBB integrity was well preserved in the combination group when compared with the lesion and standalone groups. Moreover, the combined intervention reduced the level of pro-inflammatory cytokines TNFα, NFkB, IL1α, IL1-β, IL-6, CD68, COX-2, and mRNA expression of inflammatory genes IL1α, IL1-β, IL-6, IBA-1, and COX-2 effectively. In conclusion, the present study suggests that rMCAo induced neuroinflammation and neurobehavioural alterations were attenuated by intervention with a combination of Fucoidan and cerebrolysin; Further, Fucoidan and Cerebrolysin combination improved the ischemic tolerance level by promoting the proteins and genes that regulate the inflammatory cytokines and in aiding better recovery after ischemic reperfusion injury.

The Effect of Increasing Concentrations of Omega-3 Fatty Acids from either Flaxseed Oil or Preformed Docosahexaenoic Acid on Fatty Acid Composition, Plasma Oxylipin, and Immune Response of Laying Hens

BackgroundThere is a lack of nutrition guidelines for the feeding of omega-3 polyunsaturated fatty acids (PUFA) to laying hens. Knowledge as to whether the type and concentrations of α-linolenic acid (ALA) and/or docosahexaenoic acid (DHA) in the diet can make a difference to the birds’ immune responses when subjected to a lipopolysaccharide (LPS) challenge is limited. ObjectivesThe study was designed to determine the potential nutritional and health benefits to laying hens when receiving dietary omega-3 PUFA from either ALA or DHA. MethodsA total of 80 Lohmann LSL-Classic (white egg layer, 20 wk old) were randomly assigned to 1 of 8 treatment diets (10 hens/treatment), provided 0.2%, 0.4%, 0.6%, or 0.8% of total dietary omega-3 PUFA, provided as either ALA-rich flaxseed oil or DHA-enriched algal biomass. After an 8-wk feeding period, the birds were challenged with Escherichia coli-derived LPS (8 mg/kg; i.v. injection), with terminal sample collection 4 h after challenge. Egg yolk, plasma, liver, and spleen samples were collected for subsequent analyses. ResultsIncreasing dietary omega-3 supplementation yielded predictable responses in egg yolk, plasma, and liver fatty acid concentrations. Dietary intake of ALA contributed mainly to ALA-derived oxylipins. Meanwhile, eicosapentaenoic acid- and DHA-derived oxylipins were primarily influenced by DHA dietary intake. LPS increased the concentrations of almost all the omega-6 PUFA-, ALA-, and DHA-derived oxylipins in plasma and decreased hepatic mRNA expression of COX-2 and 5-LOX (P < 0.001) involved in the biosynthesis of oxylipins. LPS also increased mRNA expression of proinflammatory cytokine IFN-γ and receptor TLR-4 (P < 0.001) in the spleen. ConclusionsThese results revealed that dietary intake of ALA and DHA had unique impacts on fatty acid deposition and their derived oxylipins and inflammatory responses under the administration of LPS in laying hens.

Long non-coding RNA reprogramming (LncRNA-ROR) attenuate inflammation induced by LPS through the regulation of miRNA-124-3p and inhibition of C5a and TLR4/Myd88/NF-κB pathwaysin C28/12 cells

Purpose: Osteoarthritis (OA), inflammation of joint and articular cartilage, is a serious joint disorder and a major cause of pain and disability. LncRNAs exert their role in OA progression. The purpose of this present research is to investigate the impact and molecular mechanism of lncRNA regulator of reprogramming (LncRNA-ROR) on inflammatory damage induced by lipopolysaccharide (LPS) in C28/12 cells. &#x0D; Methods: The expressions of lncRNA-ROR and miR-124-3p and the mRNA expressions of COX-2, iNOS, IL-8, C5a, and TNF-α were determined in C28/12 cells stimulated with LPS using RT-qPCR. Viability of C28/12 cells was evaluated using CCK-8 assay and C28/12 cell apoptosis was assessed by flow cytometry in LPS-induced C28/12 cells. Moreover, the concentration of IL-8, C5a, and TNF-α and the protein expressions of TLR4/Myd88/NF-κB pathways were examined using Elisa assay. &#x0D; Results: Results revealed that LPS inhibited the proliferation, enhanced rate of cell apoptosis, and promoted mRNA expressions of iNOS, IL-8, C5a, and TNF-α in C28/12 cells. Besides, the expression of lncRNA-ROR was expressively reduced in C28/12 cells stimulated with LPS and the lncRNA-ROR overexpression markedly reduced the inflammatory injury stimulated by LPS in C28/12 cells. Besides, lncRNA-ROR significantly down-regulated the miR-124-3p expression and miR-124-3p exposure inhibited the protecting impact of lncRNA-ROR on LPS-induced C28/12 cells. Furthermore, lncRNA-ROR suppressed the TLR4/Myd88/NF-κB pathways via suppressing the expression of miR-124-3p in LPS-induced C28/12 cells. &#x0D; Conclusion: The finding revealed that lncRNA-ROR protected against LPS-stimulated inflammatory injury in C28/12 cells by suppressing TLR4/Myd88/NF-κB pathways via inhibiting miR-124-3p suggested an efficient therapeutic strategy for OA treatment.

Functional Polymorphisms of the Arachidonic Acid Pathway Associate with Risks and Clinical Outcomes of Allergic Diseases

Introduction: The arachidonic acid (AA) pathway plays a crucial role in allergic inflammatory diseases; however, the functional roles of allergy-associated single nucleotide polymorphisms (SNPs) in this pathway remain incompletely illustrated. Methods: This study belongs to a part of an ongoing Singapore/Malaysia cross-sectional genetics and epidemiological study (SMCSGES). We performed population genotyping on n = 2,880 individuals from the SMCSGES cohort to assess the associations of SNPs in the AA pathway genes with asthma and allergic rhinitis (AR). Spirometry assessments were performed to identify associations between SNPs and lung function among n = 74 pediatric asthmatic patients from the same cohort. Allergy-associated SNPs were functionally characterized using in vitro promoter luciferase assay, along with DNA methylome and transcriptome data of n = 237 peripheral blood mononuclear cell (PBMC) samples collected from a subset of the SMCSGES cohort. Results: Genetic association analysis showed 5 tag-SNPs from 4 AA pathway genes were significantly associated with asthma (rs689466 at COX2, rs35744894 at hematopoietic PGD2 synthase (HPGDS), rs11097414 at HPGDS, rs7167 at CRTH2, and rs5758 at TBXA2R, p < 0.05), whereas 3 tag-SNPs from HPGDS (rs35744894, rs11097414, and rs11097411) and 2 tag-SNPs from PTGDR (rs8019916 and rs41312470) were significantly associated with AR (p < 0.05). The asthma-associated rs689466 regulates COX2 promoter activity and associates with COX2 mRNA expression in PBMC. The allergy-associated rs1344612 was significantly associated with poorer lung function, increased risks of asthma and AR, and increased HPGDS promoter activity. The allergy-associated rs8019916 regulates PTGDR promoter activity and DNA methylation levels of cg23022053 and cg18369034 in PBMC. The asthma-associated rs7167 affects CRTH2 expression by regulating the methylation level of cg19192256 in PBMC. Conclusions: The present study identified multiple allergy-associated SNPs that modulate the transcript expressions of key genes in the AA pathway. The development of a “personalized medicine” approach with consideration of genetic influences on the AA pathway may hopefully result in efficacious strategies to manage and treat allergic diseases.

Polydatin reduces aflatoxin-B1 induced oxidative stress, DNA damage, and inflammatory cytokine levels in mice.

This study showed the protective effect of polydatin (PD), which has an antioxidant activity against oxidative stress in mice caused by aflatoxin B1 (AFB1). In this study, 36 male Swiss albino mice were divided equally into 6 groups: 0.2mL of FTS was administered to the control group, 0.2mL of olive oil to the second group, and 0.75mg/kg AFB1 to the third group by intragastric gavage every day for 28days. The fourth, fifth, and sixth groups were administered 50, 100, and 200mg/kg PD and 0.75mg/kg AFB1 intragastrically for 28days, respectively. AFB1 administration increased plasma aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, blood urea nitrogen, creatinine, and malondialdehyde levels in blood and tissue samples but decreased the level of glutathione and the activities of superoxide dismutase and catalase. On the other hand, it was determined that PD applications depending on the increasing doses brought these levels closer to normal. In addition, AFB1 administration increased the amount of ssDNA and liver COX-2, TNF-α, IL-6, NFκB, and Cyp3a11 mRNA expression levels; on the other hand, it decreased the IL-2 mRNA expression level. In contrast, increasing doses of PD application regulated the amount of ssDNA and these mRNA expression levels. Additionally, histopathological damage was observed in the liver and kidney tissues of the AFB1 group, while PD applications in a dose-dependent manner improved these damages. As a result, it was determined that PD reduced AFB1-induced oxidative stress, DNA damage, and inflammation and exhibited a protective effect on tissues in mice.

Jinfeng pills ameliorate premature ovarian insufficiency induced by cyclophosphamide in rats and correlate to modulating IL-17A/IL-6 axis and MEK/ERK signals.

Jinfeng Pill (JFP) is a classical Chinese medicine formula and composed of 9 herbs, including Epimedium brevicornu Maxim (Yinyanghuo), Cervus elaphus Linnaeus (Lurong), Panax ginseng C.A.Mey. (Renshen), Equus asinus (EJiao), Ligustrum lucidum W.T.Aiton (Nvzhenzi), Reynoutria multiflora (Thunb.) Moldenke (Heshouwu), Curculigo orchioides Gaertn (Xianmao), Neolitsea cassia (L.) Kosterm. (Rougui) and Leonurus japonicus Houtt. (Yimucao). The formula is clinically used to regulate menstrual cycle and alleviate polycystic ovarian syndrome due to its capabilities of ovulation induction. It is therefore presumed that JFP could be used for the therapy of premature ovarian insufficiency (POI) but the assumed efficacy has not been fully substantiated in experiment. To evaluate the effectiveness of JFP on cyclophosphamide (CTX)-induced POI and preliminarily explore its potential mechanisms of action. An experimental rat model of POI was established by using CTX induction to assess the efficacy of JFP. The potential targets of action for JFP alleviating POI were predicted by the combination of network pharmacology and transcriptomics and finally validating by RT-qPCR and Western blot. JFP alleviated the damages of ovarian tissue induced by CTX in the rat model of POI via significantly decreasing serum levels of FSH and LH and the ratio of FSH/LH and increasing the levels of E2 and AMH, accompanied with promoting ovarian folliculogenesis and follicle maturity and reversing the depletion of follicle pool. With the analysis of network pharmacology, pathways in cancer, proteoglycans in cancer, PI3K-AKT, TNF and FoxO signaling pathways were predicted to be influenced by JFP. The results of RNA-seq further revealed that IL-17 signaling pathway was the most important pathway regulated by both CTX and JFP, following by transcriptional misregulation in cancer and proteoglycans in cancer. Combining the two analytical methods, JFP likely targeted genes associated with immune regulation, including COX-2, HSP90AA1, FOS, MMP3 and MAPK11 and pathways, including IL-17,Th17cell differentiation and TNF signaling pathway. Finally, JFP was validated to regulate the mRNA expression of FOS, FOSB, FOSL1, MMP3, MMP13 and COX-2 and decrease the release of IL-17A and the protein expression of IL-6 and suppress the phosphorylation of MEK1/2 and ERK1/2 in CTX induced POI rats. Jinfeng Pill is effective to ameliorate the symptoms of POI induced by CTX in the model of rats and its action is likely associated with suppressing IL-17A/IL-6 axis and the activity of MEK1/2-ERK1/2 signaling.

Study on the anti-sepsis mechanism of ursolic acid by targeting myeloid differentiation protein-2

To explore the mechanism of ursolic acid in treating sepsis using myeloid differentiation protein-2 (MD-2) as the research carrier. The affinity of ursolic acid and MD-2 was determined by biofilm interferometry technique, and the bonding mode between ursolic acid and MD-2 was tested with the aid of molecular docking technique. Raw 264.7 cells were cultured in RPMI 1640 medium and subcultured was conducted when the cell density reached 80%-90%. The second-generation cells were used for in the experiment. The effects of 8, 40 and 100 mg/L ursolic acid on cell viability were assessed by methyl thiazolyl tetrazolium (MTT) method. Cells were divided into blank group, lipopolysaccharide (LPS) group (LPS 100 μg/L) and ursolic acid group (100 μg/L LPS treatment after addition of 8, 40 or 100 mg/L ursolic acid). The effect of ursolic acid on the release of cytokines nitric oxide (NO), tumor necrosis factor-α (TNF-α) and interleukins (IL-6, IL-1β) were evaluated by enzyme-linked immunosorbent assay (ELISA). The influence of ursolic acid on the mRNA expressions of TNF-α, IL-6, IL-1β, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) were detected by reverse transcription-polymerase chain reaction (RT-PCR). The implication of ursolic acid on the protein expressions of LPS-Toll-like receptor 4 (TLR4)/MD-2-nuclear factor-κB (NF-κB) pathway were tested by Western blotting. Ursolic acid could bind to the hydrophobic cavity of MD-2 through hydrophobic bond with the amino acid residues of the protein. Therefore, ursolic acid showed high affinity with MD-2 [dissociation constant (KD) = 1.43×10-4]. The cell viability were decreased slightly, with the concentration of ursolic acid increasing, and the cell viability of 8, 40 and 100 mg/L ursolic acid were 96.01%, 94.32% and 92.12%, respectively, and there was no significant difference compared with the blank group (100%). Compared with the blank group, the cytokine level of the LPS group was significantly increased. The level of cytokines were significantly reduced by the treatment of 8, 40 and 100 mg/L ursolic acid, and the higher the concentration, the more obvious effect [compared between 100 mg/L ursolic acid group and LPS group: IL-1β (μmol/L): 38.018±0.675 vs. 111.324±1.262, IL-6 (μmol/L): 35.052±1.664 vs. 115.255±5.392, TNF-α (μmol/L): 39.078±2.741 vs. 119.035±4.269, NO (μmol/L): 40.885±2.372 vs. 123.405±1.291, all P < 0.01]. Compared with the blank group, the mRNA expressions of TNF-α, IL-6, IL-1β, iNOS and COX-2 in the LPS group were significantly increased, and the protein expressions of MD-2, myeloid differentiation factor 88 (MyD88), phosphorylation NF-κB p65 (p-NF-κB p65) and iNOS in the LPS-TLR4/MD-2-NF-κB pathway were significantly up-regulated. Compared with the LPS group, the mRNA expressions of TNF-α, IL-6, IL-1β, iNOS and COX-2 were significantly reduced by the treatment of 100 mg/L ursolic acid bound with MD-2 protein [TNF-α (2-ΔΔCt): 4.659±0.821 vs. 8.652±0.787, IL-6 (2-ΔΔCt): 4.296±0.802 vs. 11.132±1.615, IL-1β (2-ΔΔCt): 4.482±1.224 vs. 11.758±1.324, iNOS (2-ΔΔCt): 1.785±0.529 vs. 4.249±0.811, COX-2 (2-ΔΔCt): 5.591±1.586 vs. 16.953±1.651, all P < 0.01], and the proteins expressions of MD-2, MyD88, p-NF-κB p65 and iNOS in the LPS-TLR4/MD-2-NF-κB pathway were significantly down-regulated (MD-2/β-actin: 0.191±0.038 vs. 0.704±0.049, MyD88/β-actin: 0.470±0.042 vs. 0.875±0.058, p-NF-κB p65/β-actin: 0.178±0.012 vs. 0.571±0.012, iNOS/β-actin: 0.247±0.035 vs. 0.549±0.033, all P < 0.01). However, there was no difference in protein expression of NF-κB p65 among the three groups. Ursolic acid inhibits the release and expression of cytokines and mediators and regulates LPS-TLR4/MD-2-NF-κB signaling pathway by blocking MD-2 protein, and thus plays an anti-sepsis role.

Effect of Grape Seed Proanthocyanidins on Fat Metabolism and Adipocytokines in Obese Rats.

This study aimed to investigate the effect of Grape Seed Proanthocyanidin (GSP) on fat metabolism and adipocytokines in obese rats. Fifty 5-week-old rats were randomly assigned to five groups (n = 10 per group) and given either a basal diet, a high-fat diet, or a high-fat diet supplemented with GSP (25, 50, and 100 mg/d) per group. The experiment lasted for five weeks, including a one-week adaptation period and a four-week treatment period. At the end of the experimental period, serum and adipose tissue samples were collected and analyzed. Additionally, we co-cultured 3T3-L1 preadipocytes with varying concentrations of GSP to explore its effect on adipocyte metabolism. The results demonstrated that GSP supplementation reduced weight, daily gain, and abdominal fat weight coefficient (p < 0.05). It also decreased levels of glucose, cholesterol (TC) (p < 0.05), triglycerides (TG) (p < 0.05), low-density lipoprotein (LDL), cyclooxygenase-2 (COX-2), and interleukin-6 (IL-6) in adipose tissue. Furthermore, GSP addition caused adipocyte crumpling in vitro and reduced the mRNA expression of COX-2, LEP, and TNF-α in adipocytes in vitro. These findings provide compelling evidence for exploring the role of GSP in the prevention and treatment of obesity and related diseases.

Taurochenodeoxycholic acid reduces astrocytic neuroinflammation and alleviates experimental autoimmune encephalomyelitis in mice

ObjectiveMultiple sclerosis (MS) is an immune regulatory disease that affects the central nervous system (CNS). The main pathological features include demyelination and neurodegeneration, and the pathogenesis is associated with astrocytic neuroinflammation. Taurochenodeoxycholic acid (TCDCA) is one of the conjugated bile acids in animal bile, and it is not clear whether TCDCA could improve MS by inhibiting the activation of astrocytes. This study was aimed to evaluate the effects of TCDCA on experimental autoimmune encephalomyelitis (EAE)—a classical animal model of MS, and to probe its mechanism from the aspect of suppressing astrocytic neuroinflammation. It is expected to prompt the potential application of TCDCA for the treatment of MS. ResultsTCDCA effectively alleviated the progression of EAE and improved the impaired neurobehavior in mice. It mitigated the hyperactivation of astrocytes and down-regulated the mRNA expression levels of inducible nitric oxide synthase (iNOS), cyclooxygenase 2 (COX2), tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and IL-6 in the brain cortex. In the C6 astrocytic cell line induced by lipopolysaccharide (LPS), TCDCA treatment dose-dependently decreased the production of NO and the protein expression of iNOS and glial fibrillary acidic protein (GFAP). TCDCA consistently inhibited the mRNA expressions of COX2, iNOS and other inflammatory mediators. Furthermore, TCDCA decreased the protein expression of phosphorylated serine/threonine kinase (AKT), inhibitor of NFκB α (IκBα) and nuclear factor κB (NFκB). And TCDCA also inhibited the nuclear translocation of NFκB. Conversely, as an inhibitor of the G-protein coupled bile acid receptor Gpbar1 (TGR5), triamterene eliminated the effects of TCDCA in LPS-stimulated C6 cells. ConclusionTCDCA improves the progress of EAE by inhibiting the astrocytic neuroinflammation, which might be exerted by the regulation of TGR5 mediated AKT/NFκB signaling pathway. These findings may prompt the potential application of TCDCA for MS therapy by suppressing astrocyte inflammation.

Extraction optimization, structure features, and bioactivities of two polysaccharides from Corydalis decumbens.

Two polysaccharides (CPS1 and CPW2) from Corydalis decumbens were obtained to develop insights into natural medical resources. Optimal extraction conditions of total sugars were researched using the method of response surface methodology, polysaccharides were purified using a combination of ethanol precipitation and anion-exchange chromatography, and structure features were analyzed by scanning electron microscopy, transmission electron microscopy, and Congo-red assay. The bioactivities were estimated in terms of antioxidant and anti-inflammatory effects. Total sugars were extracted with an experimental yield of 32.74% under optimum conditions. CPS1 and CPW2 were purified with yields of 12.01% and 8.23%, respectively. CPS1 was a unique polysaccharide with a molecular weight (Mw) of 360 kDa and consisted of glucose, galactose, mannose, and arabinose in a ratio of 4.9:2.0:1:1.9, and CPW2 was composed of glucose with the Mw of 550 kDa. CPS1 possessed a four-helix conformation, and CPW2 was identified as a linear molecule without branched and entangled chains. The mRNA expressions of TNF-α (71.80%), IL-1β (56.55%), IL-6 (43.98%), and COX-2 (91.88%) in LPS-stimulated RAW 264.7 cells were significantly inhibited by 75 μg/mL CPS1 (P < 0.0001), while CPW2 showed lower inhibitory effects than CPS1. Compared with CPW2, CPS1 showed stronger scavenging abilities for hydroxyl (EC50 = 520.46 μg/mL), ABTS (EC50 = 533.99 μg/mL), and superoxide (EC50 = 1512.06 μg/mL) radicals. CPS1 with four-helix conformation exhibited more outstanding bioactivities than CPW2 without entangled chains.

Flavonoids in Amomum tsaoko Crevost et Lemarie Ameliorate Loperamide-Induced Constipation in Mice by Regulating Gut Microbiota and Related Metabolites.

Amomum tsaoko (AT) is a dietary botanical with laxative properties; however, the active ingredients and mechanisms are still unclear. The active fraction of AT aqueous extract (ATAE) for promoting defecation in slow transit constipation mice is the ethanol-soluble part (ATES). The total flavonoids of ATES (ATTF) were the main active component. ATTF significantly increased the abundance of Lactobacillus and Bacillus and reduced the dominant commensals, such as Lachnospiraceae, thereby changing the gut microbiota structure and composition. Meanwhile, ATTF changed the gut metabolites mainly enriched in pathways such as the serotonergic synapse. In addition, ATTF increased the serum serotonin (5-HT) content and mRNA expression of 5-hydroxytryptamine receptor 2A (5-HT2A), Phospholipase A2 (PLA2), and Cyclooxygenase-2 (COX2), which are involved in the serotonergic synaptic pathway. ATTF increased Transient receptor potential A1 (TRPA1), which promotes the release of 5-HT, and Myosin light chain 3(MLC3), which promotes smooth muscle motility. Notably, we established a network between gut microbiota, gut metabolites, and host parameters. The dominant gut microbiota Lactobacillus and Bacillus, prostaglandin J2 (PGJ2) and laxative phenotypes showed the most significant associations. The above results suggest that ATTF may relieve constipation by regulating the gut microbiota and serotonergic synaptic pathway and has great potential for laxative drug development in the future.

Low-dose nafamostat mesilate ameliorates tissue injury and inhibits 5-hydroxytryptamine synthesis in the rat intestine after methotrexate administration

We aimed to clarify the effect of nafamostat mesilate (nafamostat) on intestinal mucositis as well as the potentiation of intestinal 5-hydroxytryptamine (5-HT) dynamics induced by methotrexate, an anti-cancer drug, in rats. Rats received intraperitoneal methotrexate at 12.5 mg/kg/day for 4 days. In addition, 1, 3, or 10 mg/kg/day of nafamostat was given subcutaneously for 4 days. Ninety-six hours after the first administration of methotrexate, jejunal tissues were collected for analysis. The results showed that 1 mg/kg, but not 3 or 10 mg/kg, of nafamostat significantly ameliorated the methotrexate-induced body weight loss. Moreover, 1 mg/kg of nafamostat significantly improved methotrexate-induced mucositis, including villus atrophy. Nafamostat (1 mg/kg) significantly inhibited the methotrexate-induced mRNA expression of pro-inflammatory cytokines and cyclooxygenase-2, as well as methotrexate-induced 5-HT content and tryptophan hydroxylase (TPH) activity. In addition, it tended to inhibit the number of anti-TPH antibody-positive cells and significantly inhibited the number of anti-substance P antibody-positive cells. These findings suggest that low-dose nafamostat ameliorates tissue injury and 5-HT and substance P synthesis in methotrexate-induced mucositis. Nafamostat may be a novel therapeutic strategy for the prevention and treatment of mucositis as well as 5-HT- and/or substance P-related adverse effects in cancer chemotherapy.

Gestational low dietary protein induces intrauterine inflammation and alters the programming of adiposity and insulin sensitivity in the adult offspring

Malnutrition associated with low dietary protein can induce gestational inflammation and sets a long-lasting metabolic impact on the offspring even after replenishment. The work investigated whether a low-protein diet (LPD) during pregnancy and lactation induces intrauterine inflammation and predisposes offspring to adiposity and insulin resistance in their adult life. Female Golden Syrian hamsters were fed LPD (10.0% energy from protein) or a control diet (CD, 20.0 % energy from protein) from preconception until lactation. All pups were switched to CD after lactation and continued until the end. Maternal LPD increased intrauterine inflammation by enhancing neutrophil infiltration, amniotic hsCRP, oxidative stress, and mRNA expression of NFκβ, IL8, COX2, and TGFβ in the chorioamniotic membrane (P<.05). The prepregnancy body weight, placental, and fetal weights, serum AST and ALT were decreased, while blood platelets, lymphocytes, insulin, and HDL were significantly increased in LPD-fed dams (P<.05). A postnatal switch to an adequate protein could not prevent hyperlipidemia in the 6-months LPD/CD offspring. The lipid profile and liver functions were restored over 10 months of protein feeding but failed to normalize fasting glucose and body fat accumulation compared to CD/CD. LPD/CD showed elevated GLUT4 expression & activated pIRS1 in the skeletal muscle and increased expression of IL6, IL1β, and p65-NFκB proteins in the liver (P<.05). In conclusion, present data suggest that maternal protein restriction may induce intrauterine inflammation and affect liver inflammation in the adult offspring by an influx of fats from adipose that may alter lipid metabolism and reduce insulin sensitivity in skeletal muscle.

Anti-inflammatory activity of essential oils from Tunisian aromatic and medicinal plants and their major constituents in THP-1 macrophages

In this study, the capacity of eight essential oils (EOs), sage (Salvia officinalis), coriander (Coriandrum sativum), rosemary (Rosmarinus officinalis), black cumin (Nigella sativa), prickly juniper (Juniperus oxycedrus), geranium (Pelargonium graveolens), oregano (Origanum vulgare) and wormwood (Artemisia herba-alba), on the inhibition of NF-κB activation was screened at concentrations up to 0.25 µL/mL using THP-1 human macrophages bearing a NF-κB reporter. This screening selected coriander, geranium, and wormwood EOs as the most active, which later evidenced the ability to decrease over 50 % IL-6, IL-1β, TNF-α and COX-2 mRNA expression in LPS-stimulated THP-1 macrophages. The chemical composition of selected EOs was performed by gas chromatography–mass spectrometry (GC–MS). The two major constituents (>50 % of each EO) were tested at the same concentrations presented in each EO. It was demonstrated that the major compound or the binary mixtures of the two major compounds could explain the anti-inflammatory effects reported for the crude EOs. Additionally, the selected EOs also inhibit>50 % caspase-1 activity. However, this effect could not be attributed to the major components (except for β-citronellol/geranium oil, 40 %/65 % caspase-1 inhibition), suggesting, in addition to potential synergistic effects, the presence of minor compounds with caspase-1 inhibitory activity.These results demonstrated the potential use of the EOs obtained from Tunisian flora as valuable sources of anti-inflammatory agents providing beneficial health effects by reducing the levels of inflammatory mediators involved in the genesis of several diseases.

The Effect of Combined Treatment of Psilocybin and Eugenol on Lipopolysaccharide-Induced Brain Inflammation in Mice.

Inflammation is an organism's biological defense mechanism. Acute and chronic inflammation of the body triggers the production of pro- and anti-inflammatory pathways that can affect the content of cytokines in the brain and thus cause brain inflammation. Disorders such as depression and posttraumatic stress disorder (PTSD) are often associated with elevated inflammation. Recently, positive and promising clinical results of psilocybin for the treatment of depression and PTSD were reported. Thus, we decided to test whether psilocybin alone or in combination with eugenol, an anti-inflammatory and antioxidant agent, would prevent the increase in or decrease the content of cytokines in the brain of C57BL/6J mice injected with lipopolysaccharides (LPS). Two experiments were performed, one with pre-treatment of mice through gavage with psilocybin (0.88 mg/kg), eugenol (17.6 mg/kg), or combinations of psilocybin and eugenol (1:10, 1:20, or 1:50), followed by intraperitoneal injection of LPS, and the second, post-treatment, with initial injection with LPS, followed by treatment with psilocybin, eugenol, or their combination. Brain tissues were collected, and cytokines were analyzed by qRT-PCR, Western blot, and ELISA. Data were analyzed with a one-way ANOVA followed by Tukey's post hoc test or with multiple unpaired t-tests. LPS upregulated mRNA expression of COX-2, TNF-α, IL-1β, and IL-6. All pre-treatments decreased the expression of COX-2 and TNF-α, with psilocybin alone and in 1:50 combination, with eugenol being the most effective. In the post-treatment, all combinations of psilocybin and eugenol were effective in reducing inflammation, with the 1:50 ratio displaying the most prominent results in reducing the mRNA content of tested cytokines. Western blot analysis confirmed the effect on COX-2 and IL-1β proteins. Finally, the ELISA showed that post-treatment with psilocybin + eugenol (1:50) demonstrated the best results, decreasing the expression of multiple markers including IL-6 and IL-8. This demonstrates the anti-inflammatory effects of a combination of psilocybin and eugenol in the brain of animals with systemically induced inflammation.