Lipopolysaccharide-treated RAW Research Articles

Structural characterization and anti-inflammatory effects of polysaccharides from fruit calyx of Physalis alkekengi L. var. franchetii (Mast.) Makino

Physalis alkekengi L. (PM) is a traditional Chinese medicine with valuable nutritional and medicinal potential. Polysaccharides were considered as the important active component of calyx of P. alkekengi (PMC). In this study, four polysaccharides (PMCPs) were successfully isolated and purified by extraction, deproteination, dialysis and column chromatography. The structure characterization showed that PMCPs were mainly composed of mannose, glucosamine, ribose, rhamnose, glucose, sorbose, glucuronic acid and galacturonic acid with the molecular weights of 16.27, 71.53, 77.17 and 299.45 kDa. The FT-IR and NMR analysis suggested the presence of α and β configurations of PMCPs. PMCPs have the triple helix conformation which was determined by Congo red analysis. Moreover, the potential in vitro anti-inflammatory activity of PMCPs was investigated in lipopolysaccharide-treated RAW264.7 cells. The results indicated that PMCPs could significantly inhibit NO production at the experimental concentrations. S-PMCP-3 was found to decrease the production of pro-inflammatory cytokines including PGE2, TNF-α, IL-1β and IL-6. The findings indicated that PMCPs may be a promising natural source for addressing inflammatory damage.

Characterization and In Vitro Antioxidant and Anti-Inflammatory Activities of Ginsenosides Extracted from Forest-Grown Wild Panax quinquefolius L.

American ginseng (Panax quinquefolius L.) is known for its health benefits, which are attributed to various terpenoids. However, the specific composition and activities of these terpenoids in forest-grown wild American ginseng remain understudied. This study aimed to characterize the terpenoid composition, particularly triterpene saponins, in forest-grown wild American ginseng. The analysis revealed that triterpene saponins, notably American ginseng ginsenosides (AGGs), are the predominant active components, as identified through LC-MS/MS and HPLC. A subsequent in vitro evaluation of AGGs showcased their potent antioxidant capabilities, displaying the dose-dependent scavenging of free radicals and reducing agents. Moreover, AGGs demonstrated efficacy in reducing oxidative injury and intracellular ROS levels in RAW 264.7 macrophages treated with H2O2. In addition to their antioxidant properties, AGGs exhibited anti-inflammatory effects, significantly inhibiting NO and inflammatory substance production in lipopolysaccharide-treated RAW 264.7 macrophages. These findings highlight the potential of AGG-rich forest-grown wild American ginseng as a functional food with promising implications for improving human health.

Nuezhenoside G13 from Osmanthus fragrans fruit ameliorates Concanavalin A–induced autoimmune hepatitis by regulating the NF-κB/MAPK pathway

Ethnopharmacological relevanceOsmanthus fragrans fruit (OFF) exhibits hepatoprotective function, and it is consumed as food and used in traditional medicine in China. Nuezhenoside G13 (G13) is present in the highest levels in OFF. Autoimmune hepatitis (AIH) is a manifestation of liver disease and seriously endangers health. However, it remains unclear whether G13 affects AIH. Aim of the studyTo clarify the effect of G13 on AIH and its exact underlying mechanism from a new perspective. Materials and methodsWe used a Concanavalin A–induced AIH mouse model and lipopolysaccharide-treated Raw264.7 cells to quantify serum biochemical indicators and confirm whether G13 exhibited protective effects in the AIH mice. Furthermore, we evaluated the effect of G13 via hematoxylin and eosin and immunohistochemical staining. We used enzyme-linked immunosorbent assay (ELISA) and polymerase chain reaction to quantify the inflammatory factors. We confirmed that G13 inhibited apoptosis via terminal deoxynucleotidyl transferase dUTP nick end labeling staining. Molecular docking, immunofluorescence, and western blotting experiments of G13 and key proteins of the NF-κB/MAPK pathway revealed that G13 alleviated inflammation. In addition, Cell Counting Kit-8, ELISA, NO detection, and western blotting assays were performed. Finally, we used an inhibitor of the p38 MAPK to verify that G13 reduced inflammation through the NF-κB/MAPK pathway in Raw264.7 cells. ResultsThe in vivo experiments revealed that G13 improved oxidative stress and apoptosis. In addition, G13 decreased the expression levels of CD4+, CD8+, F4/80+, and Ly6G and the secretion of inflammatory factors. Interestingly, G13 reduced the phosphorylation levels of IκBα, NF-κB, JNK, ERK1/2, and p38. Additionally, the in vitro experiments revealed that G13 alleviated inflammation through the NF-κB/MAPK pathway in lipopolysaccharide-treated Raw264.7 cells. Furthermore, molecular docking demonstrated that the binding fraction of G13 with these proteins was high. ConclusionG13 suppressed oxidative stress, apoptosis, and inflammation in a Concanavalin A–induced AIH mouse model. Furthermore, G13 exerted its effect through the NF-κB/MAPK pathway.

Puerarin Inhibits NLRP3-Caspase-1-GSDMD-Mediated Pyroptosis via P2X7 Receptor in Cardiomyocytes and Macrophages.

Diabetic cardiomyopathy (DCM) is a critical complication of long-term chronic diabetes mellitus, and it is characterized by myocardial fibrosis and myocardial hypertrophy. Previous studies have shown that the pyroptosis pathway was significantly activated in DCM and may be related to the P2X7 receptor. However, the role of the P2X7 receptor in the development of DCM with pyroptosis is still unclear. In this study, we aimed to explore the mechanism of puerarin and whether the P2X7 receptor can be used as a new target for puerarin in the treatment of DCM. We adopted systematic pharmacology and bioinformatic approaches to identify the potential targets of puerarin for treating DCM. Additionally, we employed D-glucose-induced H9C2 rat cardiomyocytes and lipopolysaccharide-treated RAW264.7 mouse mononuclear macrophages as the in vitro model on DCM research, which is close to the pathological conditions. The mRNA expression of cytokines in H9C2 cells and RAW264.7 macrophages was detected. The protein expressions of NLRP3, N-GSDMD, cleaved-caspase-1, and the P2X7 receptor were investigated with Western blot analysis. Furthermore, molecular docking of puerarin and the P2X7 receptor was conducted based on CDOCKER. A total of 348 puerarin targets and 4556 diabetic cardiomyopathy targets were detected, of which 218 were cross targets. We demonstrated that puerarin is effective in enhancing cardiomyocyte viability and improving mitochondrial function. In addition, puerarin is efficacious in blocking NLRP3-Caspase-1-GSDMD-mediated pyroptosis in H9C2 cells and RAW264.7 cells, alleviating cellular inflammation. On the other hand, similar experimental results were obtained by intervention with the P2X7 receptor antagonist A740003, suggesting that the protective effects of puerarin are related to the P2X7 receptor. The molecular docking results indicated key binding activity between the P2X7 receptor and puerarin. These findings indicate that puerarin effectively regulated the pyroptosis signaling pathway during DCM, and this regulation was associated with the P2X7 receptor.

An MMP-2 Responsive Nanotheranostic Probe Enabled Synergistic Therapy of Rheumatoid Arthritis and MR/CT Assessment of Therapeutic Response In Situ.

This study reports a facile and green synthesis of a new multifunctional nanotheranostic probe for the synergistic therapy of rheumatoid arthritis (RA) and in situ assessment of therapeutic response. The probe is synthesized through a one-step self-assembly of two exquisitely designed peptide-amphiphilic block copolymers (PEG-DTIPA-KGPLGVRK-MTX and Pal-GGGGHHHHD-TCZ) under mild conditions, requiring minimal energy input. The resultant probe demonstrates excellent biocompatibility, water solubility, and colloidal stability. It exhibits a strong IL-6R targeting ability toward inflamed joints, and releases drugs in an MMP-2-responsive manner. The co-loading of methotrexate(MTX) and tocilizumab (TCZ) into the probe enables synergistic RA therapy with improved efficacy by simultaneously decreasing the activity of adenosine synthetase and interfering with the binding of IL-6 to its receptor. In addition, the resultant probe exhibits a high r1 relaxation rate (7.00 mm-1 s-1 ) and X-ray absorption capability (69.04 Hu mm-1 ), enabling sensitive MR and CT dual-modal imaging for simultaneous evaluation of synovial thickness and bone erosion. Both in vitro experiments using lipopolysaccharide-treated RAW264.7 cells and in vivo experiments using collagen-induced arthritis mice demonstrate the probe's high effectiveness in synergistically inhibiting inflammation. This study provides new insights into RA theranostics, therapeutic monitoring, the design of multifunctional theranostic probes, and beyond.

Structural characterization and anti-inflammatory activities of anti-complementary polysaccharides from Rhododendron principis.

Rhododendron principis leaves have been used as "Dama", a Traditional Tibetan Medicine for treating inflammatory diseases. R. principis crude polysaccharides (CZND) with anti-complementary activity demonstrated promising anti-inflammatory effects on acute lung injury (ALI) induced by lipopolysaccharide (LPS). CZND significantly decreased the levels of TNF-α and IL-6 in both serum and BALF (blood and bronchoalveolar lavage fluid) in LPS-induced ALI mice by intragastric administration (100 mg/kg). A heteropolysaccharide, ZNDHP, was obtained from CZND with successive anti-complementary activity-guided separation. ZNDHP was characterized as a branched neutral polysaccharide with a backbone composed of →2)-β-Glcp-(1→, →2,6)-α-Glcp-(1→, →6,3)-β-Galp-(1→, →2,6)-α-Galp-(1→, →6,2)-β-Glcp-(1→, →4)-α-Glcp-(1→, →5)-β-Araf-(1→, →3,5)-α-Araf-(1→, and →4,6)-β-Manp-(1→, and the backbone structure was further confirmed by partial acid hydrolysis. In addition to anti-complementary and antioxidant activities, ZNDHP exhibited potent anti-inflammatory activity by significantly inhibiting the secretion of NO, TNF-α, IL-6, and IL-1β of LPS treated RAW264.7 cells. However, all of these activities decreased greatly after partially hydrolyzing, indicating the importance of the multi-branched structure for its bioactivity. Therefore, ZNDHP might be an important component of R. principis for treating inflammation.

Hyptis obtusiflora C. Presl ex Benth Methanolic Extract Exhibits Anti-Inflammatory and Anti-Gastritis Activities via Suppressing AKT/NF-κB Pathway

Inflammation is an indispensable part of the human body's self-defense mechanism against external stimuli. The interactions between Toll-like receptors and microbial components trigger the innate immune system via NF-κB signaling, which regulates the overall cell signaling including inflammatory responses and immune modulations. The anti-inflammatory effects of Hyptis obtusiflora C. Presl ex Benth, which has been used as a home remedy for gastrointestinal disorders and skin disease in rural areas of Latin America, have not yet been studied. Here, we investigate the medicinal properties of Hyptis obtusiflora C. Presl ex Benth methanol extract (Ho-ME) for inflammatory response suppression. Nitric oxide secretion in RAW264.7 cells triggered by TLR2, 3, or 4 agonists was reduced by Ho-ME. Reduction of inducible nitric oxide synthase (iNOS), cyclooxygenase (COX)-2, and interleukin (IL)-1b mRNA expression was observed. Decreased transcriptional activity in TRIF- and MyD88-overexpressing HEK293T cells was detected with a luciferase assay. Additionally, serially downregulated phosphorylation of kinase in the NF-κB pathway by Ho-ME was discovered in lipopolysaccharide-treated RAW264.7 cells. Together with the overexpression of its constructs, AKT was identified as a target protein of Ho-ME, and its binding domains were reaffirmed. Moreover, Ho-ME exerted gastroprotective effects in an acute gastritis mouse model generated by the administration of HCl and EtOH. In conclusion, Ho-ME downregulates inflammation via AKT targeting in the NF-κB pathway, and the combined results support Hyptis obtusiflora as a new candidate anti-inflammatory drug.

Ferroptosis Inhibitors Suppress Prostaglandin Synthesis in Lipopolysaccharide-Stimulated Macrophages.

Necrostatin-1 blocks ferroptosis via an unknown mechanism and necroptosis through inhibition of receptor-interacting protein kinase-1 (RIP1). We report that necrostatin-1 suppresses cyclooxygenase-2-dependent prostaglandin biosynthesis in lipopolysaccharide-treated RAW264.7 macrophages (IC50 ∼ 100 μM). This activity is shared by necrostatin-1i (IC50 ∼ 50 μM), which lacks RIP1 inhibitory activity, but not the RIP1 inhibitors necrostatin-1s or deschloronecrostatin-1s. Furthermore, we show that the potent ferroptosis inhibitors and related compounds ferrostatin-1, phenoxazine, phenothiazine, and 10-methylphenothiazine strongly inhibit cellular prostaglandin biosynthesis with IC50's in the range of 30 nM to 3.5 μM. None of the compounds inhibit lipopolysaccharide-mediated cyclooxygenase-2 protein induction. In the presence of activating hydroperoxides, the necrostatins and ferroptosis inhibitors range from low potency inhibition to stimulation of in vitro cyclooxygenase-2 activity; however, inhibitory potency is increased under conditions of low peroxide tone. The ferroptosis inhibitors are highly effective reducing substrates for cyclooxygenase-2's peroxidase activity, suggesting that they act by suppressing hydroperoxide-mediated activation of the cyclooxygenase active site. In contrast, for the necrostatins, cellular prostaglandin synthesis inhibition does not correlate with peroxidase-reducing activity but rather with the presence of a thiohydantoin substituent, which conveys the ability to reduce the endoperoxide intermediate prostaglandin H2 to prostaglandin F2α in vitro. This finding suggests that necrostatin-1 blocks cellular prostaglandin synthesis and ferroptosis via a redox mechanism distinct from action as a one-electron donor. The results indicate that a wide range of compounds derived from redox-active chemical scaffolds can block cellular prostaglandin biosynthesis.

Antioxidant and anti-inflammatory effects of solar salt brined kimchi.

Salt is an essential ingredient in the kimchi fermentation process. Solar salt has antioxidant, anti-cancer, and anti-obesity properties. The aim of this study was to determine the antioxidant and anti-inflammatory effects of solar salt brined kimchi. Purified salt (PS), dehydrated solar salt (DSS), 1-year aged solar salt (SS1), and 3-years aged solar salt (SS3) were investigated. Anti-inflammatory effects were determined by analyzing cytotoxicity, nitric oxide (NO) production, and inflammation-related gene expression in lipopolysaccharide-treated RAW264.7 cells. Antioxidant activities of DSS, SS1, and SS3 were higher than that of PS. Solar salt significantly inhibited NO production with low cytotoxicity and decreased inflammation-related gene expression. Kimchi containing solar salt (DSSK, SS1K, and SS3K) showed higher antioxidant activity than PSK. Additionally, DSSK, SS1K, and SS3K significantly inhibited NO production and decreased the expression of inflammation-related genes. Owing to the antioxidant and anti-inflammatory effects, using solar salt in kimchi preparation could have potential health benefits.

MiR-222-3p Aggravates the Inflammatory Response by Targeting SOCS1 to Activate STAT3 Signaling in Ulcerative Colitis.

Ulcerative colitis is characterized by relapsing inflammation in the gastrointestinal tract with limited treatment options. The aim of the present study was to assess the anti-inflammatory effect of Suppressor of cytokine signaling (SOCS1) on lipopolysac- charide-stimulated RAW264.7 cells and to investigate its potential mechanisms. The in vitro ulcerative colitis model was established by using lipopolysaccharide-stimulated RAW264.7 cells. Western blot- ting was used to detect the protein expression levels of SOCS1, JAK2, STAT3, and VDR. Reverse transcription-quantitative polymerase chain reaction was used to measure the mRNA expression of SOCS1, miR-222-3p, and VDR. An enzyme-linked immunosorbent assay was performed to measure the levels of inflammatory cytokines. A luciferase assay assessed the binding of SOCS1 to miR-222-3p. A total of 15 patients with ulcerative colitis and 18 healthy controls were recruited. The expression levels of SOCS1 and miR-222-3p in the colonic mucosa tissues of patients with ulcerative colitis and healthy controls were determined by reverse transcription-quantitative polymerase chain reaction. SOCS1 upregulation inhibited the lipopolysaccharide-induced inflammation in RAW264.7 cells. SOCS1 was confirmed to be tar- geted by miR-222-3p. Silencing SOCS1 significantly abolished the inhibitory effects of miR-222-3p downregulation on inflammation. MiR-222-3p activated STAT3 signaling and reduced VDR expression by targeting SOCS1 in lipopolysaccharide-treated RAW264.7 cells. Additionally, miR-222-3p expression was upregulated in ulcerative colitis patients (P = 5.16E-10), while SOCS1 (P = 2.75E-10) and VDR (P = 52.5E-9) expression was downregulated in ulcerative colitis patients. Endoscopic scores (UCEIS) revealed significant positive cor- relation with miR-222-3p and negative correlation with SOCS1 and VDR. MiR-222-3p targets SOCS1 to aggravate the inflammatory response by suppressing VDR and activating STAT3 signaling in ulcerative colitis.

Phenolic compounds from Chaenomeles speciosa alleviate inflammation in lipopolysaccharide-treated RAW264.7 macrophages via the NF-κB and MAPK pathways

Chaenomeles speciosa (Sweet) Nakai cultivated widely in temperate regions possesses anti-inflammatory properties, however, the underlying molecular mechanisms remain not fully understood. In this study, a purified phenolic extract of C. speciosa rich in chlorogenic acid, procyanidin B1 and catechin (determined by HPLC-Q-TOF-MS/MS) exhibited dose-dependent anti-inflammatory effects on lipopolysaccharide (LPS)-treated RAW264.7 macrophages. The extract at 30 μg/mL was most potent and enabled most cells in normal morphology under LPS stimulation without causing cytotoxicity. The extract suppressed the levels of nitric oxide (NO), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1β (IL-1β), and the mRNA and protein expressions of nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). The mechanisms underlying such anti-inflammatory actions included the regulation of phosphorylation of related proteins to monitor the expressions of inflammatory mediators and genes in the nuclear factor kappa-B (NF-κB) and mitogen-activated protein kinase (MAPK) signaling pathways. Therefore, the phenolic extract from C. speciosa is a desirable anti-inflammatory agent for inflammatory conditions to meet the rising demand for natural and cost-effective therapeutics.

Anti-inflammatory effects of Torin2 on lipopolysaccharide-treated RAW264.7 murine macrophages and potential mechanisms

ContextTorin2 has various pharmacological properties. However, its anti-inflammatory activity has not been reported. ObjectiveThis study focused on the potential anti-inflammatory properties of Torin2 in lipopolysaccharide (LPS)-evoked RAW264.7 murine macrophages. The study aimed to shed light on the molecular mechanisms that ameliorate these effects. MethodsTorin2 was applied to 100 ng/mL lipopolysaccharide-induced RAW 264.7 macrophages in vitro. Nitric oxide (NO) levels were detected using the Griess reagent kit. Prostaglandin E2 (PGE2), pro-inflammatory cytokines interleukin (IL)-1β, interleukin (IL)-6, and tumor necrosis factor in the supernatant fraction were determined using enzyme-linked immunosorbent assay (ELISA). Gene expression of pro-inflammatory cytokines, cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS) were tested using real-time quantitative polymerase chain reaction (qPCR). Cyclooxygenase-2 and inducible nitric oxide synthase proteins, phosphorylation of mitogen-activated protein kinase (MAPK) subgroups, extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), p38, I-kappa-B-alpha (IκBα), and nuclear factor-kappa-B (NF-κB), and activation in extracts were detected via western blotting. Nuclear factor-kappa-B/p65 nuclear translocation was tested using an immunofluorescence assay. ResultsThe results demonstrated that pre-treatment with Torin2 profoundly attenuated the lipopolysaccharide-stimulated levels of nitric oxide and prostaglandin E2, pro-inflammatory cytokines, messenger ribonucleic acid (mRNA), and protein expression of cyclooxygenase-2 and inducible nitric oxide synthase. Collectively, Torin2 pre-treatment notably weakened lipopolysaccharide-induced damage by reducing the phosphorylation of nuclear factor-kappa-B, p38, c-Jun N-terminal kinase, extracellular signal-regulated kinase proteins, and nuclear factor-kappa-B/p65 nuclear translocation. ConclusionNumerous pieces of evidence indicated that Torin2 reversed inflammatory activation by regulating nuclear factor-kappa-B and mitogen-activated protein kinase signaling pathways and provided a tentative potential candidate for preventing and treating inflammatory diseases.

Fuzhuan brick tea polysaccharides serve as a promising candidate for remodeling the gut microbiota from colitis subjects in vitro: Fermentation characteristic and anti-inflammatory activity

The purified fraction 3 of polysaccharides from Fuzhuan brick tea (FBTPS-3) could attenuate the colitis and modulate the gut microbiota. However, the relationship between anti-inflammatory effect of FBTPS-3 and the gut microbiota is still unknown. Thus, the anaerobic fermentation in vitro was used to investigate the potential mechanism. FBTPS-3 could be utilized and degraded by gut microbiota from inflammatory bowel disease (IBD) subjects. Furthermore, FBTPS-3 could modulate the composition and structure of IBD gut microbiota toward to that of healthy group. FBTPS-3 showed a superior modulated effect on IBD gut microbiota by increasing Bacteroides and decreasing Escherichia/Shigella. Furthermore, the fermentation solution rather than FBTPS-3 itself showed anti-inflammatory effects on lipopolysaccharide-treated RAW264.7 macrophages, which might be due to the metabolites such as short-chain fatty acids (SCFAs). Thus, FBTPS-3 can be expected as novel prebiotics for treatment of IBD via modulating gut microbiota, and promoting the production of SCFAs.

Four New Highly Oxygenated Eremophilane Sesquiterpenes from an Endophytic Fungus Boeremia exigua Isolated from Fritillaria hupehensis.

Four new eremophilane-type sesquiterpenes, boeremialanes A–D (1–4) were obtained from solid substrate cultures of Boeremia exigua (Didymellaceae), an endophytic fungus isolated from Fritillaria hupehensis (Liliaceae). Boeremialanes A–C (1–3) are highly oxygenated eremophilanes with a benzoate unit attached at the C-13 position and are rarely found in nature. Their structures and absolute configurations were determined by extensive spectroscopic methods, electronic circular dichroism (ECD), and nuclear magnetic resonance (NMR) calculations with DP4+ analysis. Boeremialane D (4) potently inhibited nitric oxide production in lipopolysaccharide-treated RAW264.7 macrophages with an IC50 of 8.62 μM and was more potent than the positive control, pyrrolidinedithiocarbamate (IC50 = 23.1 μM).

Characterization and Evaluation of Antioxidant and Anti-Inflammatory Activities of Flavonoids from the Fruits of Lycium barbarum.

The fruits of Lycium barbarum are rich in flavonoids, which may contribute to the health-promoting function of Lycium barbarum. However, the composition of flavonoids in the fruits of Lycium barbarum (LBFs) has received little attention. Thus, the goal of this work was to identify more kinds of flavonoids from fruits of Lycium barbarum by liquid chromatography–mass spectrometry. The potential antioxidant and anti-inflammatory activities of LBFs in vitro were also investigated. Thirteen flavonoid compounds were identified in LBFs, of which daphnetin, 6,7-dihydroxycoumarin, astragalin, taxifolin, eriodictyol, naringenin, and chrysoeriol were identified for the first time in the fruits of Lycium barbarum, which greatly enriched the variety of flavonoids in the fruits of Lycium barbarum. LBFs showed a similar superior antioxidant activity to vitamin C. Furthermore, LBFs exhibited an anti-inflammatory activity by suppressing the production of nitric oxide and pro-inflammatory cytokines, including tumor necrosis factor-alpha, interleukin-1β, and interleukin-6, in lipopolysaccharide-treated RAW264.7 macrophage cells. This study demonstrated the potential development of LBFs as functional foods.

Qualitative and quantitative analyses of aconite alkaloids in Aconiti kusnezoffii Radix, and NO inhibitory activity evaluation of the alkaloid extracts

Aconiti kusnezoffii Radix (Caowu), a traditional Chinese medicine and Mongolian medicine, has been commonly used owing to its analgesic and anti-inflammatory effects. Here, we performed a qualitative analysis of aconite alkaloids (AAs) in raw and processed Aconiti kusnezoffii Radix by ultra-high-performance liquid chromatography (HPLC) electrospray ionization-tandem mass spectrometry. Furthermore, we quantified benzoylaconine, benzoylmesaconine, benzoylhypaconine, aconitine, mesaconitine, and hypaconitine in raw and processed Aconiti kusnezoffii Radix using a HPLC system coupled with a diode array detector. The analysis of AA fragmentation patterns in positive electrospray ionization mode revealed 155 AAs, including 6 with unambiguous identities, 109 with putative assignments, and 40 unknown compounds. Additionally, lipopolysaccharide-treated RAW264.7 macrophages were used to evaluate the anti-inflammatory properties of the alkaloid extracts of Aconiti kusnezoffii Radix. Importantly, alkaloid extracts of Aconiti kusnezoffii Radix at three different concentrations exhibited protective effect against lipopolysaccharide-induced inflammation. Thus, the proposed method can be used not only for the qualitative and quantitative analyses of Aconiti kusnezoffii Radix, but also for evaluating other Aconitum spp. Our study lays a foundation for the clinical application of poisonous Aconitum.

Alkyl-benzofuran dimers from Eupatorium chinense with insulin-sensitizing and anti-inflammatory activities

Five new racemic alkyl-benzofuran dimers, (±)-dieupachinins I–M (1–5), were isolated from the root tubers of Eupatorium chinense, a well-known traditional Chinese medicine for the treatment of diphtheria in Guangdong province. The structures of these compounds, especially the first examples of 12,10′-epoxy dimer dieupachinin I (1), 12-nor-dimer dieupachinin J (2), and 12,12′-dinor-dimer dieupachinin K (3), were elucidated by spectroscopic data analysis. Chiral resolution were further carried out on a cellulose column by HPLC, and compounds 2–5 were successfully separated into two enantiomers, respectively. The absolute configurations of (+)-(2–5) and (−)-(2–5) were established by theoretical ECD calculation. All the compounds were evaluated for insulin-stimulated glucose uptake in C2C12 myotubes and (±)-dieupachinin I (1) exhibited the best activity. Compound 1 enhanced insulin-stimulated glucose uptake via activating the insulin receptor substrate 1/protein kinase B/glycogen synthase kinase-3β signaling pathway. Moreover, all the isolates were tested for their nitric oxygen (NO) inhibitory effects in lipopolysaccharide-treated RAW264.7 macrophages, and compounds (±)-1, (±)-2, and (±)-4 showed promising inhibitory effects with IC50 values of 6.42 ± 1.85, 6.29 ± 1.94, and 16.03 ± 2.07 μM, respectively. (±)-Dieupachinin I (1) again dose-dependently suppressed LPS-induced expression of inducible NO synthase and nuclear translocation of p65.

BIOACTIVE SAPONINS FROM PANAX BIPINNATIFIDUS SEEM. GROWING IN VIETNAM

Panax bipinnatifidus Seem. belonging to the family Araliaceae has been used as a folk medicine in Viet Nam. Our ongoing study on bioactive compounds from Panax bipinnatifidus Seem. collected in Sapa, Lao Cai resulted in the isolation of two saponins together with the well-known saponin Stipuleanoside R2. Their structures were identified as 3-O-β-D-glucopyranosyl-(1→3)-β-D-glucuronopyranosyloleanolic acid (1) and Stipuleanoside R1 (2) based on NMR spectroscopic data as well as comparison with the literature data. For the first time, 3-O-β-D-glucopyranosyl-(1→3)-β-D-glucuronopyranosyloleanolic acid (1) was found in genus Panax. On biological screening, the crude extract and the principal saponin, stipuleanoside R2 showed weak inhibitory effect on markedly nitric oxide production in lipopolysaccharide-treated RAW 264.7 cells.

Isorhapontigenin alleviates lipopolysaccharide-induced acute lung injury via modulating Nrf2 signaling

Nuclear factor erythroid-2 related factor 2 (Nrf2) is involved in mitigating various oxidative stress- and inflammation-induced diseases, including acute lung injury/acute respiratory distress syndrome (ALI/ARDS). Isorhapontigenin (ISO), from the Chinese herb Gnetum cleistostachyum, exhibits antioxidant and anti-inflammatory properties. In this study, we explored the protective effects of ISO in ALI and its underlying molecular mechanisms. ISO significantly mitigated ALI by reducing the lung wet/dry weight ratio, protein concentration in the bronchoalveolar lavage fluid (BALF), and the levels of myeloperoxidase and malondialdehyde. ISO also improved the superoxide dismutase and glutathione activity in vivo. Moreover, ISO effectively ameliorated the changes in IL-1β, IL-6, and TNF-α concentrations in BALF, prevented IκB degradation, and inhibited the phosphorylation of NF-κB p65 subunit in lung tissues; furthermore, it enhanced the nuclear translocation of Nrf2 and inhibited IL-1β, IL-6, TNF-α, iNOS, COX-2, and ROS production in lipopolysaccharide-treated RAW264.7 cells. The protective effects of ISO in ALI were significantly reversed in ML385-treated RAW264.7 cells and the mouse model, indicating its role in Nrf2-activation. In conclusion, ISO effectively ameliorated lipopolysaccharide-induced ALI by reducing inflammation and oxidative stress, primarily through activation of Nrf2 signaling.

Isolation and Characterization of New Anti-Inflammatory and Antioxidant Components from Deep Marine-Derived Fungus Myrothecium SP. Bzo-l062.

In the present study, four new compounds including a pair of 2-benzoyl tetrahydrofuran enantiomers, namely, (−)-1S-myrothecol (1a) and (+)-1R-myrothecol (1b), a methoxy-myrothecol racemate (2), and an azaphilone derivative, myrothin (3), were isolated along with four known compounds (4–7) from cultures of the deep-sea fungus Myrothecium sp. BZO-L062. Enantiomeric compounds 1a and 1b were separated through normal-phase chiral high-performance liquid chromatography. The absolute configurations of 1a, 1b, and 3 were assigned by ECD spectra. Among them, the new compound 1a and its enantiomer 1b exhibited anti-inflammatory activity, inhibited nitric oxide formation in lipopolysaccharide-treated RAW264.7 cells, and exhibited antioxidant activity in the 2,2-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) and oxygen radical absorbance capacity assays.