LPS-induced NO Production In RAW264 Research Articles

Seven New Noroleanane Triterpenoids from Mastic and their NO Inhibitory and Cytotoxic Activities.

Mastic is a natural resin produced by Pistacia lentiscus L. (Anacardiaceae) with high medicinal value and have been traditionally used as Uighur imported medicine for centuries. In this study, 16 triterpenoids including seven new norleananetriterpenoids (1-7), along with nine known oleanane triterpenoids (8-16), were isolated from the mastic. Their chemical structures were determined on the basis of extensive spectroscopic analyses (including IR, UV, ESI-HR-MS and NMR spectroscopy) and single-crystal X-ray diffraction. Compounds 4-7, 11, 14 and 16 showed strong inhibitory NO productionin LPS-induced RAW264.7 cells with IC50 values 7.44-9.76 μM, respectively (positive control dexamethasone, 9.93 ± 1.17 μM). Furthermore,compounds 3 and 12 significantly inhibited the growth of SW480 cells, compound 3 showed the most pronounced inhibitory effect with an IC50 of 2.30 ± 0.38 μM.

Sesquiterpenoids from Carpesium abrotanoides and their anti-inflammatory activity both in vitro and in vivo

Twenty-nine sesquiterpenoids, including pseudoguaiane-type (1–11), eudesmane-type (12–23), and carabrane-type (24–29), have been identified from the plant Carpesium abrotanoides. Of them, compounds 1–4, 12–15, and 24–27, namely carpabrotins A–L, are twelve previously undescribed ones. Compound 3 possessed a pseudoguaiane backbone with a rearrangement modification at C-11, C-12 and C-13, while compound 4 suffered a carbon bond break between the C-4 and C-5 to form a rare 4,5-seco-pseudoguaiane lactone. Compounds 1–3, 5, 13–16 and 25–27 exhibited anti-inflammatory activity by inhibiting NO production in LPS-induced RAW264.7 macrophages with IC50 values less than 40 μM, while compounds 1, 2, 5, 13, 14, 16, and 25–27 showed significant inhibitory activity comparable to that of dexamethasone. The anti-atopic dermatitis (AD) effects of compounds 5 and 16 were tested according to 2,4-dinitrochlorobenzene (DNCB)-induced AD-like skin lesions in KM mice, and the results revealed that the major products 5 and 16 improved the histological features of AD-like skin lesions and mast cell infiltration in mice This study suggested that sesquiterpenoids in C. abrotanoides should play a key role in its anti-inflammatory use.

Millingtonia hortensis L.f. ethanol extract exerts in vivo and in vitro anti-inflammatory activities through inhibition of Syk kinase in NF-κB pathway

Ethnopharmacological relevanceMillingtonia hortensis L.f., commonly known as tree jasmine or Indian cork tree, is native to South Asia and Southeast Asia. Traditionally, its stem bark, leaves, and roots are employed for pulmonary, gastrointestinal, and antimicrobial purposes, while the flowers are used in treating asthma and sinusitis.Aim of the study: The underlying anti-inflammatory mechanisms of M. hortensis remain relatively unexplored. Therefore, we studied the anti-inflammatory effects of M. hortensis and the molecular mechanisms of its ethanol extracts (Mh-EE) both in vitro and in vivo. Materials and methodsNitric oxide (NO) production was assessed using Griess reagent, while cell viability of RAW264.7 cell and HEK293T cells were determined via the MTT assay. Constituent analysis of Mh-EE using GC/MS-MS and HPLC, and mRNA expression of inflammatory cytokines was measured through PCR and RT-PCR. Protein levels were analyzed using western blotting. The thermal stability of Mh-EE was evaluated by CESTA. Lastly, a gastritis in vivo model was induced by HCl/EtOH, and protein expression levels were measured using western blotting. ResultsMh-EE significantly reduced NO production in LPS-induced RAW264.7 cells without substantially affecting cell viability. Additionally, Mh-EE decreased the expression of proinflammatory factors, such as iNOS, IL-1β and COX2. Furthermore, Mh-EE downregulated TLR4 expression, altered MyD88 recruitment, and suppressed phosphorylation of Syk, IKKα, IκBα and AKT. Simultaneously, Mh-EE also attenuated NF-κB signaling in HCl/EtOH-induced mice. ConclusionsMh-EE exerts anti-inflammatory effects by suppressing p-Syk in the NF-κB pathway, and it has potential as a novel treatment agent for inflammatory diseases.

Glycoside constituents from Cayratia geniculata

Using various chromatographic separations, six glycoside derivatives (1–6), including one new ent-labdane glucoside named cayratioside (1), were isolated from the methanol extract of Cayratia geniculata stems and leaves. Their structures were elucidated by detailed analysis of the 1D, 2D NMR, and HRESIQTOF mass spectra. The inhibitory effect of 1–6 on LPS-induced NO production in RAW264.7 cells was also evaluated. Among isolated compounds, 1 exhibited moderate activity with an IC50 value of 59.65 ± 1.85 µM.

Anti-inflammatory lindenane sesquiterpenoid dimers from the roots of Chloranthus holostegius var. trichoneurus.

Two new lindenane-type sesquiterpenoid dimers, chlotrichenes C and D (1 and 2) together with five known lindenane-type sesquiterpenoid dimers (3-7) were isolated from the roots of Chloranthus holostegius var. trichoneurus, a famous natural medicine named as "Sikuaiwa" for subduing swellings and relieving pain. The structures including absolute configuration were elucidated by their 1D and 2D NMR, HRESIMS, and ECD data. Compounds 1 and 2 were classical [4 + 2] lindenane-type sesquiterpenoid dimers that differed from known analogs in oxidation profile, side chain profile, and double bond position. The new isolates and compound 3 exhibited significant inhibitory activity on IL-1β production (IC50: 1-15μM) in LPS-induced THP-1 cells and other compounds exhibited inhibitory activity on NO production in LPS-induced RAW 264.7 cells (IC50: 24-33μM).

A metallothionein from disk abalone (Haliotis discus discus): Insights into its functional roles in immune response, metal tolerance, and oxidative stress

Metallothioneins (MTs) are cysteine-rich metal-binding proteins whose expression is induced by exposure to essential and non-essential metals, making them potential biological markers for assessing metal pollution in various biomonitoring programs. However, the functional properties of these proteins are yet to be comprehensively characterized in most marine invertebrates. In this study, we identified and characterized an MT homolog from the disk abalone (Haliotis discus discus), referred to as disk abalone MT (AbMT). AbMT exhibited the same primary structural features as MTs from other mollusks containing two β-domains (β2β1-form). AbMT protein demonstrated metal-binding and detoxification abilities against Zn, Cu, and Cd, as evidenced by Escherichia coli growth kinetics, metal tolerance analysis, and UV absorption spectrum. Transcriptional analysis revealed that AbMT was ubiquitously expressed in all analyzed tissues and upregulated in gill tissue following challenge with Vibrio parahaemolyticus, Listeria monocytogenes, and viral hemorrhagic septicemia virus (VHSV). Additionally, overexpression of AbMT suppressed LPS-induced NO production in RAW264.7 macrophages, protected cells against H2O2-induced oxidative stress, and promoted macrophage polarization toward the M1 phase. Conclusively, these findings suggest an important role for AbMT in environmental stress protection and immune regulation in disk abalone.

Diterpenoid glucosides with anti-inflammatory activity from Sigesbeckia glabrescens

Six previously undescribed diterpenoid glucosides, along with four known compounds, were isolated from the aerial parts of Sigesbeckia glabrescens. The structures and absolute configurations of undescribed compounds were elucidated using extensive spectroscopic techniques, ECD calculations and chemical methods. Compounds 1 and 8 exhibited anti-inflammatory activity against LPS-induced NO production in RAW 264.7 macrophages, with compound 8 demonstrating significant inhibitory activity compared to positive control minocycline, boasting an IC50 value at 14.20 μM.

Two new megastigmane glycosides from the leaves of Nicotiana tabacum and their anti-inflammatory activities

Two new megastigmane glycosides, (6 R,7E,9R)-3-oxo-α-ionyl-9-O-α-L-rhamnopyranosyl-(1′′→4′)-β-D-glucopyranoside (1) and (6 R,7E,9R)-3-oxo-α-ionyl-9-O-β-D-glucopyranosyl-(1′′→6′)-β-D-glucopyranoside (2), together with six known analogues (3–8) were isolated from the leaves of Nicotiana tabacum. The structures of all metabolites were determined by comprehensive analysis of NMR and MS spectroscopic data as well as by comparison with those of previously reported. The in vitro anti-inflammatory activity of all isolates was evaluated using a lipopolysaccharide (LPS)-induced RAW264.7 cell inflammatory model, and the compounds 1, 3, 7, and 8 exhibited inhibition of LPS-induced NO production in RAW264.7 macrophage cells with IC50 values of 42.3-61.7 μM (positive control, dexamethasone, IC50 = 21.3 ± 1.2 μM).

Coumarins and flavones with anti-inflammatory activity isolated from the branches and leaves of Daphne retusa.

In this study, two new (1, 13) and fourteen known (2-12, 14-16) compounds were isolated from the branches and leaves of Daphne retusa. On the basis of chemical evidence and spectral data analysis (UV, ECD NMR, and HR-ESI-MS), the structures of new compounds were elucidated. Furthermore, all compounds have been tested for their inhibitory effects on NO production in LPS-induced RAW 264.7 cells, and compound 3 showed obvious inhibitory effect. Through target screening and molecular docking technology, potential binding targets for compound 3 to exert anti-inflammatory effects have been predicted.

Bioactive cytochalasans from the desert soil-derived fungus Chaetomium madrasense 375 obtained via a chemical engineering strategy.

The chemical engineering of natural extracts has emerged as an effective strategy for the production of diverse libraries of chemicals, making it integral to drug discovery. A chemical engineering strategy based on the epoxidation and ring-opening reactions was used to prepare diversity-enhanced extracts of Chaetomium madrasense 375. Eleven unnatural cytochalasan derivatives (1-11) with unique functional groups, such as amine and isoxazole, were isolated and characterized from these chemically engineered extracts of C. madrasense 375. The identification of these new structures was accomplished through comprehensive spectroscopic analysis, supplemented by synthetic considerations. Notably, compounds 5 and 13-16 displayed potent phytotoxic effects on Arabidopsis thaliana, while compounds 1, 2, 5, 10, and 12 demonstrated inhibitory activities on LPS-induced NO production in RAW264.7 cells. Among them, compound 1 was found to be able to inhibit the upregulated expression of the inducible nitric oxide synthase (iNOS) protein induced by LPS, while also decreasing the production of pro-inflammatory cytokines (IL-6) and influencing the phosphorylation of p38, ERK1/2, and JNK at 100 μM. Our findings demonstrate that the chemical engineering of natural product extracts can be an efficient technique for the generation of novel bioactive molecules.

Dimeric ent-kauranoids isolated from Isodon japonica var. Glaucocalyx and their anti-inflammatory activities

The phytochemical investigation of the aerial parts of Isodon japonica var. glaucocalyx afforded four undescribed (glaucocalyxin O-R, 1–4) and six known ent-kauranoids (5–10). Their structures were established using NMR and MS measurements. Compounds 1 and 2 are dimeric ent-kaurane-type diterpenoids. Moreover, the plausible biogenetic pathways for compounds 1 and 2 were proposed as Michael addition between two monomers. Eight compounds were assayed for their anti-inflammatory activity by evaluating NO production in LPS-induced RAW 267.4 cells, and compounds 7, 8 and 9 exhibited relatively remarkable anti-inflammatory activities at 10 μM.

Triterpenoids in Sorbus pohuashanensis suspension cell treated with yeast extract

This study induced biological stress in Sorbus pohuashanensis suspension cell(SPSC) with yeast extract(YE) as a bio-tic elicitor and isolated and identified secondary metabolites of triterpenoids produced under stress conditions. Twenty-six triterpenoids, including fifteen ursane-type triterpenoids(1-15), two 18,19-seco-ursane-type triterpenoids(16-17), four lupine-type triterpenoids(18-21), two cycloartane-type triterpenoids(22-23), and three squalene-type triterpenoids(24-26), were isolated and purified from the methanol extract of SPSC by chromatography on silica gel, MCI, Sephadex LH-20, and MPLC. Their structures were elucidated by spectroscopic analyses. All triterpenoids were isolated from SPSC for the first time and 22-O-acetyltripterygic acid A(1) was identified as a new compound. Selected compounds were evaluated for antifungal, antitumor, and anti-inflammatory activities, and compound 1 showed an inhibitory effect on NO production in LPS-induced RAW264.7 cells.

Rutaecarpine analogues with potent anti-inflammation to alleviate acute ulcerative colitis via regulating TLR4/MAPK/NF-κB pathway

Natural products are a rich resource in drug discovery. In the report, series novel rutaecarpine-derived compounds with potent anti-inflammatory activity and therapeutic efficacy against dextran sulfate sodium (DSS)-induced ulcerative colitis (UC) were designed and synthesized. Thirty-two target compounds inhibiting on LPS-induced NO production in RAW264.7 cells were screened. Compound 7mA with the most potent inhibitory activity and low cytotoxicity was selected as representative compound for further studies on the anti-inflammatory efficacy and mechanism. The results indicated that 7mA could significantly inhibit the transcription or expression of iNOS, IL-6 and IL-1β via regulating the TLR4/MAPK/NF-κB signal pathway. What's more, 7mA also could alleviate UC in DSS-induced mice, and down-regulated the release of IL-6, IL-1β, inhibited activation of TLR4/MAPK/NF-κB pathway in mice model as well, suggesting that this new rutaecarpine-derived scaffold might serve as therapeutic candidate for further development to anti-inflammatory agent.

Minor protopanaxadiol type sapogenins from the alkali hydrolysate of stems-leaves of Panax notoginseng

Two new protopanaxadiol type sapogenins, (3β,12β)-3,12,20-trihydroxydammar-24-en-26-al (1) and (3β,12β)-3,12,20-trihydroxydammar-24-en-26-oic acid (2), were isolated from the alkali hydrolysate of stems-leaves of Panax notoginseng, along with seven known analogues (3–9). Their structures were elucidated by spectroscopic analyses and single-crystal X-ray diffraction. Compound 2 and the known sapogenins 5–8 displayed weak to moderate inhibition of NO production in LPS-induced RAW264.7 macrophages with IC50 values from 44.5 to 143.6 μM, respectively.

Phenolic glycosides from Yua thomsonii

Phytochemical study of Yua thomsonii aerial parts led to the isolation of seven phenolic glycosides, including three new compounds, namely yuaphenosides A−C (1−3). The structural elucidation was confirmed by a detailed analysis of the 1D and 2D NMR and HR-ESI-QTOF mass spectra. Compounds 4−7 were isolated from Y. thomsonii for the first time. Among the isolated compounds, yuaphenoside A (1) and piceatannol-3′-O-β-D-glucopyranoside (6) exhibited significant inhibitory effect on LPS-induced NO production in RAW264.7 cells. Moderate effect was observed for yuaphenoside B (2), saccharumoside B (4), piceid (5), and piceatannol-4′-O-β-D-glucopyranoside (7); whereas weak activity was found for yuaphenoside C (3).

Cytospotones A-D, four new polyketones from the endophytic fungus Cytospora sp. A879

Three new α-pyrone derivatives cytospotones A-C (1–3) and a new cyclohexenone derivative cytospotone D (4) together with four known α-pyrones were isolated from the endophytic fungus Cytospora sp. A879 of Pogostemon cablin (Blanco) Benth. The structures of 1–4 were elucidated primarily by spectroscopic methods (1D, 2D NMR and HRESIMS), ECD spectra analyses, and ECD calculations. Furthermore, the four new compounds (1–4) were evaluated for their anti-inflammatory and α-glucosidase inhibitory activities. The results showed that compound 1 had moderate inhibitory effect on LPS-induced NO production in RAW 264.7 macrophages.

Essential oils from Acronychia pedunculata (L.) Miq. in Vietnam: chemical composition, antimicrobial, cytotoxicity and nitric oxide inhibition activities

The chemical composition, antimicrobial, cytotoxic and anti-inflammatory activities of essential oils from different parts of Acronychia pedunculata, a medicinal plant native to Central Highlands of Vietnam, were investigated. Their chemical constituents were determined using GC-MS. The antimicrobial properties were assessed using a microdilution broth method, while the anti-inflammatory properties were measured based on their capacity to inhibit NO production in macrophage cells. Their cytotoxicity was tested using the sulforhodamine B assay on three human cancer cell lines. The GC-MS analysis revealed the presence mainly of caryophyllene oxide (35.3%) and spathulenol (31.9%) in the stem essential oil, caryophyllene oxide (51.3%), and linalool (21.0%) in the leaf essential oil, and α-pinene (21.4%), (E)-β-ocimene (16.5%), linalool (12.5%), and β-caryophyllene (13.0%) in the fruit essential oil. All three essential oils inhibited the growth of various bacteria and fungi, reduced NO production in LPS-induced RAW264.7 cells, and exhibited cytotoxicity against all three tested cancer cell lines. Notably, the fruit essential oil showed the most potent cytotoxicity among the essential oils, with IC50 values of 5.47 ± 0.21, 7.95 ± 0.35, and 9.22 ± 0.52 g/mL against SK-LU-1, MCF-7, and HepG2 cells, respectively. It also had a strong anti-inflammatory effect that was comparable to that of dexamethasone, with an IC50 value of NO production inhibition of 14.62 ± 0.62 mg/mL. These results suggest the potential of the essential oils from this plant to provide leads for the development of anti-inflammatory and anti-cancer agents.

Synthesis, Characterization, and in vitro Anti-Inflammatory Activity of Novel Ferrocenyl(Piperazine-1-Yl)Methanone-based Derivatives.

Inflammation is closely related to the occurrence and development of various diseases in the clinical scope. Finding effective anti-inflammatory agents is of great significance for clinical treatment. A series of novel ferrocenyl(piperazine-1-yl)methanone-based sulfamides and carboxamides were synthesized to discover potent anti-inflammatory agents. The compounds were characterized by 1H NMR, 13C NMR, and MS spectra. Compound 5h was further determined by single crystal X-ray diffraction. All the target compounds were screened for anti-inflammatory activity by evaluating the inhibition effect of LPS-induced NO production in RAW264.7 macrophages. The novel compound (4i) is the preliminary anti-inflammatory mechanism detected by western blot. In a multi-stage screening campaign, compound 4i was shortlisted, which exhibited physicochemical properties suitable for human administration. Among them, compound 4i was found to be most potent in inhibiting NO production (IC50 = 7.65 μM) with low toxicity. This compound also exhibited significant inhibition of the production of iNOS and COX-2. Preliminary mechanism studies indicated that compound 4i could inhibit the activation of the LPS-induced TLR4/NF-κB signaling pathway. The promising anti-inflammatory activity of compound 4i compared with the reference drug suggests that this compound may contribute as a lead compound in the search for new potential anti-inflammatory agents.

Dichapetalin-type triterpenoids from Dichapetalum longipetalum and their anti-inflammatory activity

A phytochemical research on the twigs of Dichapetalum longipetalum (Turcz.) Engl. Resulted in five undescribed dichapetalin-type triterpenoids 1–5. Their chemical structures were determined by spectroscopic analysis of HR-ESIMS and NMR spectra and the absolute configuration of compound 1 was completely elucidated by single crystal X-ray crystallography. Through preliminary anti-inflammatory activity assessment, compound 1 exhibited inhibitory effect on LPS-induced NO production in RAW264.7 murine macrophages with an IC50 value of 2.09 μM.

Acylated flavonoid glycosides from Barringtonia pendula and their inhibition on lipopolysaccharide-induced nitric oxide production in RAW264.7 cells

Six new acylated flavonoid glycosides namely barringosides J − O (1–6) along with tephrokaempferoside and barringoside D were isolated from the branches and leaves of Barringtonia pendula. The structural elucidation was confirmed by extensive analysis of their spectroscopic data including HRQTOFMS, 1D and 2D NMR experiments. Moderate inhibitory effects on LPS-induced NO production in RAW264.7 cells were observed for barringosides M (4) and N (5) with IC50 values of 48.40 ± 3.01 and 56.61 ± 3.87 μM, whereas weak inhibition was found for compounds 1–3, 6, and 7 with IC50 values ranging from 64.91 ± 3.68 to 79.80 ± 3.90 μM.