Nauclea officinalis inhibits inflammation in LPS-mediated RAW 264.7 macrophages by suppressing the NF-κB signaling pathway.

Nauclea officinalis has been traditionally used in China for the treatment of fever, pneumonia and enteritidis etc. This study aims to investigate effects of N. officinalis on the inflammatory response as well as the possible molecular mechanism in LPS-stimulated RAW 264.7 murine macrophage cells. Anti-inflammatory activity of N. officinalis (10, 20, 50, and 100µg/mL) was investigated by using LPS-induced RAW 264.7 macrophages. The NO production was determined by assaying nitrite in culture supernatants with the Griess reagent. The levels of TNF-α, IL-6 and IL-1β in culture media were measured with ELISA kits. Real time fluorescence quantitative PCR was detected for mRNA expression of iNOS, TNF-α, IL-6 and IL-1β. Western blot assay was performed to illustrate the inhibitory effects of N. officinalis on phosphorylation of IκB-α and NF-κB p65. Treatment with N. officinalis (10-100µg/mL) dose-dependently inhibited the production as well as mRNA expression of NO, TNF-α, IL-6 and IL-1β in RAW 264.7 macrophages. Western blot assay suggested that the mechanism of the anti-inflammatory effect was associated with the inhibition of phosphorylation of IκB-α and NF-κB p65. The results indicated that N. officinalis potentially inhibited the activation of upstream mediator NF-κB signaling pathway via suppressing phosphorylation of IκB-α and NF-κB p65 to inhibit LPS-stimulated inflammation.

Zhai XT ，Zhang ZY ，Jiang CH ，Chen JQ ，Ye JQ ，Jia XB ，Yang Y ，Ni Q ，Wang SX ，Song J ，Zhu FX ... -  《-》

Anti-inflammatory effect of the six compounds isolated from Nauclea officinalis Pierrc ex Pitard, and molecular mechanism of strictosamide via suppressing the NF-κB and MAPK signaling pathway in LPS-induced RAW 264.7 macrophages.

Nauclea officinalis Pierrc ex Pitard. is a Chinese medicinal herb that contains high level of alkaloids which is the most abundant and active constituent. Strictosamide isolated from Nauclea officinalis Pierrc ex Pitard. showed significant effects on inflammatory response, compared with pumiloside, 3-epi-pumiloside, vincosamide, 3α,5α-tetrahydrodeoxycordifoline lactam and naucleamide A-10-O-β-D-glucopyranoside of this plant. we investigated the biological activities of the six compounds mentioned-above, and the underlying molecular mechanism exerted by the most potent one, strictosamide. The effects of strictosamide and other five compounds on the inhibitory activity of nitric oxide (NO) were screened by Griess test. The contents of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) in media were detected by using Enzyme-linked immunosorbent (ELISA) kits. The effects on the mRNA expression of nitric oxide synthase (iNOS), TNF-α and IL-1β of strictosamide were further investigated by RT-qPCR. Western blot assay was conducted to illustrate the effects of strictosamide on iNOS and phosphorylation of p65, inhibitor of NF-κB (IκB)-α, IκB-kinase (IKK)-α as well as p-extracellular signal-regulated kinase (ERK), p-c-jun N-terminal kinase (JNK) and p-p38 in the protein levels. Strictosamide potently suppressed the productions of NO, TNF-α and IL-1β in LPS-induced RAW 264.7 macrophages, and it dose-dependently alleviated the LPS-simulated protein level of iNOS as well as the mRNA expressions of iNOS, TNF-α and IL-1β. In addition, molecular data revealed that strictosamide markedly decreased the expressions of p-p65, p-IκBα and p-IKKα. Furthermore, strictosamide significantly attenuated LPS-induced the phosphorylation of p38, ERK and JNK. At present study, the results indicated that the anti-inflammatory activity of strictosamide was associated with the restraint of NO, TNF-α and IL-1β via negative regulation of both NF-κB and mitogen-activated protein kinases (MAPKs) in LPS-induced RAW 264.7 cells.

Li D ，Chen J ，Ye J ，Zhai X ，Song J ，Jiang C ，Wang J ，Zhang H ，Jia X ，Zhu F ... -  《-》

DXXK exerts anti-inflammatory effects by inhibiting the lipopolysaccharide-induced NF-κB/COX-2 signalling pathway and the expression of inflammatory mediators.

Diao Xin Xue Kang (DXXK) is the active pharmaceutical ingredient of the traditional Chinese medicinal product DXXK capsules, which have been approved for the treatment of cardiovascular disease and have been widely used clinically in China for many years with distinct curative effects. In March 2012, DXXK capsules were approved in the Netherlands, making them the first traditional herbal medicinal product (THMP) made outside of Europe. To assess the anti-inflammatory effects of DXXK and the underlying mechanisms at the cellular and molecular levels. In this study, lipopolysaccharide (LPS) was used to induce inflammation in RAW 264.7 cells. The sulphorhodamine B (SRB) assay was used to study the effect of DXXK on the proliferation of RAW 264.7 cells. Gene expression levels of cyclooxygenase 1 (COX-1), COX-2, tumour necrosis factor-alpha (TNF-α), IL-1β and IL-6 were assessed by reverse transcription polymerase chain reaction (RT-PCR), while COX-2 protein levels were evaluated using western blotting. The levels of PGE2 in the culture media were detected by enzyme-linked immunosorbent assay (ELISA), while TNF-α, IL-1β and IL-6 levels were detected using a Milliplex Map Mouse Cytokine Panel system. The activation and nuclear translocation of nuclear transcription factor κB (NF-κB) were studied using western blotting. In vivo studies in mice were carried out using the carrageenan-air pouch models of inflammation. In exudates, leucocytes were counted, total protein was determined using the Bradford assay, nitric oxide(NO) levels were assessed using the Griess reagent, and PGE2 and TNF-α levels were quantified by ELISA. The SRB assay showed that at the doses used in this study (10, 20 and 40 μg/mL), DXXK did not affect the proliferation of RAW 264.7 cells. DXXK (10, 20 and 40 μg/mL) inhibited LPS-induced PGE2 production by down-regulating the expression of COX-2, without influencing COX-1 expression. We also demonstrated that DXXK reduced the expression of pro-inflammatory cytokines, including TNF-α, IL-1β and IL-6, at both the gene and protein levels. Furthermore, DXXK inhibited LPS-induced NF-κB activation and nuclear translocation by suppressing the phosphorylation of IκB. Consistent with the in vitro results, the in vivo studies demonstrated that DXXK reduced leucocyte counts as well as total protein, NO, PGE2 and TNF-α levels in the exudates of mice with carrageenan-air pouch inflammation. The current study revealed that DXXK has a significant anti-inflammatory effect that may be attributed to its inhibitory effect on the NF-κB/COX-2 pathway and associated inflammatory mediators, including PGE2, NO, TNF-α, IL-1β and IL-6. The current study provides additional evidence of the effects of DXXK in inflammation. Based on the combination of our results and previously reported data, we propose that DXXK has multiple pharmacological effects that could be harnessed to treat systemic diseases.

Yu Y ，Li X ，Qu L ，Chen Y ，Dai Y ，Wang M ，Zou W ... -  《-》

Corydalis bungeana Turcz. attenuates LPS-induced inflammatory responses via the suppression of NF-κB signaling pathway in vitro and in vivo.

Corydalis bungeana Turcz. (C. bungeana) is one of traditionally used medicines in China and possesses various biological effects, such as anti-inflammatory, antibacterial activity and inhibition of the immune function of the host. we studied the anti-inflammatory effect and molecular mechanism involved of C. bungeana both in vitro and in vivo model system in which the inflammatory response was induced by LPS treatment. Anti-inflammatory activity of C. bungeana was investigated by LPS-induced RAW 264.7 macrophages and BALB/c mice. The production and expression of pro-inflammatory cytokines were evaluated by Griess reagent, ELISA kits and RT-qPCR, respectively. Phosphorylation status of IκBα and p65 was illustrated by western blot assay. C. bungeana reduced the secretion of NO, TNF-α, IL-6 and IL-1β through inhibiting the protein expression of iNOS, TNF-α, IL-6 and IL-1β in vitro and in vivo. Western blot analysis suggested that C. bungeana supressed NF-κB activation via regulating the phosphorylation of IκBα and p65. Immunohistochemical assay also demostrated the histological inflammatory change in liver tissue. The results indicate that C. bungeana supresses the activation of NF-κB signaling pathway through inhibiting phosphorylation of IκBα and p65, which results in good anti-inflammatory effect. In addition, C. bungeana attenuates inflammatory reaction by supressing the expression of various inflammatory cytokines both in vivo and in vitro.

Zhai XT ，Chen JQ ，Jiang CH ，Song J ，Li DY ，Zhang H ，Jia XB ，Tan W ，Wang SX ，Yang Y ，Zhu FX ... -  《-》

Anti-inflammatory and antitumoural effects of Uncaria guianensis bark.

Uncaria guianensis (Aublet) Gmell (Rubiaceae) is a medicinal plant from the jungles of South and Central America, used to treat cancer, arthritis, diabetes, and inflammation. Evaluate the anti-inflammatory and anti-tumor effects of Uncaria guianensis preparations. Bio-guided fractionation of a hydroethanolic extract of Uncaria guianensis was performed, evaluating the fractions and subfractions for their effect on inflammatory mediators, tumour necrosis factor alpha (TNF-α), interleukin 6 (IL-6) and prostaglandin E2 (PGE2) by ELISA and nitric oxide (NO) by the Griess reaction in cultured supernatant from RAW 264.7 macrophages stimulated with lipopolysaccharide (LPS). The expression of cyclooxygenase 2 (COX-2), inducible nitric oxide synthase (iNOS) and inhibitor of κB (IκB) were investigated in RAW 264.7 macrophages by flow cytometry. The activity of NF-κB in HeLa cells transfected with a luciferase reporter system was determined. The effect of Uncaria guianensis on the inflammatory response in vivo was assessed in BALB/c mice stimulated with LPS, on rat paw oedema induced by carrageenan, and on tumour growth and lung metastasis in BALB/c mice inoculated with 4T1 mammary tumour cells. Immune cell infiltrates and inflammatory mediators were evaluated in the tumour by immunohistochemistry. Sub-fraction Ug AIV inhibited, to varying degrees, NO, TNF-α, IL-6 and PGE2 production by macrophages in vitro (30 μg/ml) and in the serum of LPS-challenged mice (5 mg/kg). Macrophage expression of Cox-2 was inhibited (35%), IκB degradation was completely inhibited and NF-κB activation was inhibited (70%) by Ug AIV at 30 μg/ml. Ug AIV decreased paw oedema by 86% (5 mg/kg) and serum NO and TNF-α by 45% and 65% respectively. Ug AIV reduced 4T1 mammary tumour growth by 91% on day 33 post-inoculation as well as the levels of serum NO, IL-6 and TNF-α in the same animals. Ug AIV decreased the number of tumour-infiltrating T lymphocytes, macrophages and neutrophils as well as the number of cells positive for COX-2, iNOS, IL-6, TNF-α and p65. As Ug AIV was not cytotoxic for tumour cells or macrophages, its anti-tumour effect may be due to a reduction in pro-tumoural inflammatory processes in the tumour microenvironment, possibly mediated through NF-κB.

Urdanibia I ，Michelangeli F ，Ruiz MC ，Milano B ，Taylor P ... -  《-》