CCL5 increases lung cancer migration via PI3K, Akt and NF-κB pathways

CCL5 (previously called RANTES) is in the CC-chemokine family and plays a crucial role in the migration and metastasis of human cancer cells. Besides, integrins are the major adhesive molecules in mammalian cells. Here we found CCL5 increased the migration and cell surface expression of αvβ3 integrin in human lung cancer cells (A549 cells). CCL5 stimulation increased phosphorylation of the p85α subunit of phosphatidylinositol 3-kinase (PI3K) and serine 473 of Akt. Also, we found that PI3K inhibitor (Ly294002) or Akt inhibitor suppressed CCL5-induced migration activities and integrin expression of A549 cells. Transfection of cells with p85 or Akt mutant also reduced CCL5-mediated cancer migration. In addition, treatment of A549 cells with CCL5 induced IκB kinase α/β (IKK α/β) phosphorylation, IκB phosphorylation, p65 Ser⁵³⁶ phosphorylation, and κB-luciferase activity. Furthermore, the CCL5-mediated increases in p65 Ser⁵³⁶ phosphorylation were inhibited by Ly294002 and Akt inhibitor. Taken together, our results suggest that CCL5 acts through PI3K/Akt, which in turn activates IKKα/β and NF-κB, resulting in the activation of αvβ3 integrin and contributing to the migration of human lung cancer cells.     

The aromatic ketone 4'-hydroxychalcone inhibits TNFα-induced NF-κB activation via proteasome inhibition

Chalcones are aromatic ketones, known to exhibit anti-microbial, anti-inflammatory and anti-cancer activities. The aim of this study was to investigate the anti-inflammatory and anti-cancer activity of 4′-hydroxychalcone. Here, we report that 4′-hydroxychalcone inhibits TNFα-induced NF-κB pathway activation in a dose-dependent manner. To investigate the underlying molecular mechanisms we demonstrate that 4′-hydroxychalcone inhibits proteasome activity in a dose-dependent manner but has no effect on IKK activity. Results show that 4′-hydroxychalcone inhibits TNFα-dependent degradation of IκBα and subsequently prevents p50/p65 nuclear translocation leading to 4′-hydroxychalcone-inhibited expression of NF-κB target genes. Most importantly, inhibition of NF-κB activation by 4′-hydroxychalcone is not leukemia cell-type specific and has no significant effect on non-transformed cell viability, thus highlighting the compound's potential in both prevention and treatment.     

Salvianolic acid A suppress lipopolysaccharide-induced NF-κB signaling pathway by targeting IKKβ

Salvianolic acid A, an active compound present in Salvia miltiorrhiza, is a phenolic carboxylic acid derivative, ((2R)-3-(3,4-Dihydroxyphenyl)-2-[(E)-3-[2-[(E)-2-(3,4-dihydroxyphenyl) ethenyl]-3,4-dihydroxyphenyl] prop-2-enoyl]oxypropanoic acid). The present study was performed to investigate the underlying mechanisms of anti-inflammatory effects with salvianolic acid A, specially focused on nuclear factor κB (NF-κB) signaling pathway by targeting the IκB kinase β (IKKβ). The effect of salvianolic acid A for IKKβ activity was analyzed using an immobilized metal affinity for phosphochemicals (IMAP)-based time-resolved fluorescence resonance energy transfer (TR-FRET) assay. The underlying mechanisms of salvianolic acid A were examined using lipopolysaccharide (LPS)-stimulated RAW264.7 cells. IKKβ studies based on IMAP-TR-FRET showed that salvianolic acid A possesses a potent IKKβ inhibitory activity with Ki value of 3.63 μM in an ATP-noncompetitive manner. Pretreatment with salvianolic acid A (10, 30 μM) decreased LPS-induced expression of iNOS and COX-2, thereby inhibiting production of nitric oxide and prostaglandin E₂, respectively. In addition, salvianolic acid A (10, 30 μM) also attenuated the LPS-induced IκBα phosphorylation and degradation, and NF-κB translocation. These results suggest that salvianolic acid A modulates NF-κB-dependent inflammatory pathways through IKKβ inhibition and these anti-inflammatory effects will aid in understanding the pharmacology and mode of action of salvianolic acid A.     

The inhibition of TNF-α-induced NF-κB activation by marine natural products

The deregulated activation of NF-κB is associated with cancer development and inflammatory diseases. With an aim to find new NF-κB inhibitors, we purified and characterized compounds from extracts of the Fijian sponge Rhabdastrella globostellata, the crinoid Comanthus parvicirrus, the soft corals Sarcophyton sp. nov. and Sinularia sp., and the gorgonian Subergorgia sp. after an initial screening of 266 extracts from different marine origins.Results obtained show that selected purified compounds had a cytotoxic effect on the human leukaemia cell line K562, inhibited both TNF-α-induced NF-κB-DNA binding as well as TNF-α-induced IκBα degradation and nuclear translocation of p50/p65. Furthermore, we observed the inhibition of NF-κB activation induced by an overexpression of IKKβ. Interestingly, natural products inhibited IKKβ kinase as well as the 26S proteasome proteolytic activity.     

N-乙酰半胱氨酸防治内毒素所致肝损伤的实验研究

目的　探讨N 乙酰半胱氨酸 (NAC)可否对内毒素性肝损伤进行抑制及相应的细胞、分子机制。方法　健康雄性昆明种小鼠 30只随机分为肝损伤组、NAC组和对照组 ,不同给药后检测肝匀浆肿瘤坏死因子(TNF) α、丙二醛 (MDA)和还原型谷胱甘肽 (GSH)含量变化 ,并行聚丙烯酰胺凝胶电泳 ,分析NAC对核因子(NF) κBp6 5、IκBα的影响。 结果　NAC不但使肝组织TNF α、MDA含量降低 ,GSH含量升高 ,且抑制了内毒素诱导胞质IκBα降解和NF κBp6 5的表达。 结论　NAC可能通过调整库普弗细胞氧化还原平衡 ,影响NF κBp6 5活化 (核易位 ) ,从而抑制了TNF α等炎性因子基因的表达 ,减轻肝损伤。     

阿托伐他汀对LPS诱导人血管内皮细胞IκBα表达的影响

目的通过观察阿托伐他汀对LPS诱导的人血管内皮细胞IκBα表达的影响来探明其抑制核因子κB活性的机制。方法将培养的人血管内皮细胞株ECV304,分为对照组、LPS组、阿托伐他汀低、中、高3个浓度组。阿托伐他汀3组加入不同阿托伐他汀孵育后与LPS组均加入LPS刺激30min。用免疫荧光法观察核因子κB活化情况,用逆转录-聚合酶链反应观察IκBα mRNA的表达情况,用蛋白印迹法观察IκBα及p-IκBα蛋白含量的变化。结果阿托伐他汀3组能够抑制p65由胞质转移至核内、能够剂量依赖性的降低pIκBα蛋白表达、减少IκBα的降解;阿托伐他汀高浓度组IκBα mRNA的表达增加。结论阿托伐他汀可以通过影响IκBα的表达和降解来抑制核因子κB活性。     

The molecular mechanism of polygalasaponin F-mediated decreases in TNFα: emphasizing the role of the TLR4-PI3K/AKT-NF-κB pathway

Polygalasaponin F (PS-F), an oleanane-type triterpenoid saponin extracted from Polygala japonica, decreases the release of the inflammatory cytokine tumor necrosis factor α (TNFα), but the precise molecular mechanisms by which this event occurs are not fully understood. To study the anti-neuroinflammatory mechanisms of PS-F, enzyme-linked immunosorbent assay was used to detect the secretion of TNFα from BV-2 microglial cells. Nuclear proteins extracted from BV-2 microglial cells stimulated by lipopolysaccharide (LPS) and pretreated with/without inhibitors were measured by Western blotting, and cell viability was evaluated by MTT analysis. The results indicated that inhibition of toll-like receptor (TLR) 4 (CLI-095 1 μg/ml), phosphatidylinositol 3-kinase (PI3K) (Ly294002 10 μM) or IκBα phosphorylation (Bay11-7082 10 μM) completely prevents the release of TNFα induced by LPS without affecting cell viability and attenuated the nuclear translocation of p65 stimulated by LPS. In addition, PS-F exhibited a similar trend regarding TNFα release, AKT phosphorylation and NF-κB translocation. These results suggest that PS-F reduces neuroinflammatory cytokine secretion through the regulation of the TLR4-PI3K/AKT-NF-κB signaling pathway.     

SC-514, a selective inhibitor of IKKβ attenuates RANKL-induced osteoclastogenesis and NF-κB activation

The RANKL-induced NF-κB signaling pathway is essential for osteoclastogenesis. This study aims to identify specific inhibitors targeting NF-κB signaling pathway, which might serve as useful small molecule inhibitors for the treatment and alleviation of osteoclast-mediated bone lytic diseases. By screening for compounds that selectively inhibit RANKL-induced NF-κB activation in RAW264.7 cells as monitored by luciferase reporter gene assay, we identified SC-514, a specific inhibitor of IKKβ, as a candidate compound targeting osteoclastogenesis. SC-514 dose-dependently inhibits RANKL-induced osteoclastogenesis with an IC50 of <5 μM. At high concentrations, SC-514 (≥12.5 μM) induced apoptosis and caspase 3 activation in RAW264.7 cells. Moreover, SC-514 specifically suppressed NF-κB activity owing to delayed RANKL-induced degradation of IκBα and inhibition of p65 nuclear translocation. Taken together, our results indicate that SC-514 impairs RANKL-induced osteoclastogenesis and NF-κB activation. Thus, targeting IKKβ by SC-514 presents as a potential treatment for osteoclast-related disorders such as osteoporosis and cancer-induced bone loss.     

泻心汤有效组分不同配伍对内毒素肺损伤大鼠NF-κB及IκB表达的影响

目的研究泻心汤有效组分及其配伍对内毒素肺损伤大鼠NF-κB及IκB表达的影响。方法大鼠灌胃给予黄芩总黄酮提取物(TFL)、大黄总游离蒽醌提取物(TFA)、大黄总结合蒽醌提取物(TCA)、TFL与TFA配伍高、低剂量(A高A低)、TFL与TCA配伍(B)及醋酸地塞米松(Dex)4d,末次给药后1h股静脉注射脂多糖(LPS)建立大鼠内毒素肺损伤模型,1、2、4h各组分别处死6只动物,收集肺组织,免疫印记(Western blot)检测核因子κB(NF-κB)、κB抑制因子(IκB)的蛋白表达。结果大黄总游离蒽醌、A高及Dex在1、2、4h均能够明显抑制NF-κB p65的核转位(P<0.01),所有给药组在2、4h均能够明显抑制胞质中的IκBα的降解(P<0.01)。结论泻心汤有效组分及其配伍对内毒素肺损伤大鼠保护作用与抑制NF-κB的核转位及IκB的降解有关。