铁观音茶提取物对脂多糖诱导RAW264.7细胞炎症反应的抑制作用及机制

研究铁观音茶提取物对脂多糖(LPS)诱导的RAW264.7细胞炎症反应的抑制作用及机制。用脂多糖作用于RAW264.7细胞,建立炎症模型,并用吲哚美辛和不同浓度铁观音提取物处理,检测NO和IL-6的分泌情况,qPCR检测一氧化氮合酶(iNOS)、环氧合酶2(COX-2)、肿瘤坏死因子α(TNF-α)、单核细胞趋化蛋白1(MCP-1)、白细胞介素6(IL-6)mRNA相对表达,Western Blot检测炎症相关蛋白激酶(IKKβ),核转录因子κB抑制因子(IκB)、核转录因子κB p65(NF κB p65)及其磷酸化产物的相对表达。结果显示,铁观音茶提取物能显著抑制炎症介质NO分泌和IL-6蛋白表达量(p<0.05),抑制炎症相关基因iNOS、COX-2、TNF-α和MCP-1等表达,并极显著抑制NF-κB信号通路相关蛋白IKKβ、IkB和p65的磷酸化(p<0.01)。以上结果表明,铁观音茶提取物可明显抑制LPS诱导的RAW264.7细胞炎症反应,其机制可能与抑制NF-κB信号通路激活有关。     

羊栖菜多酚通过核转录因子-κB/丝裂原活化蛋白激酶通路缓解脂多糖诱导的RAW264.7细胞炎症反应

目的：研究羊栖菜多酚对脂多糖（lipopolysaccharide,LPS）诱导小鼠单核巨噬细胞白血病细胞RAW264.7细胞炎症反应的影响。方法：噻唑蓝法测定细胞活力;Griess法测定一氧化氮（NO）水平;实时荧光定量聚合酶链式反应测定白细胞介素（interleukin,IL）-6、IL-1β、肿瘤坏死因子（tumor necrosis factor,TNF）-α、环氧合酶2（cyclooxygenase-2,COX-2）和诱导型一氧化氮合酶（inducible nitric oxide synthase,iNOS）的基因相对表达量;流式细胞术测定细胞吞噬能力;蛋白免疫印迹法测定信号通路丝裂原活化蛋白激酶（mitogen-activated protein kinases,MAPKs）和核转录因子（nuclear factor,NF）-κB信号通路关键蛋白表达水平。结果：羊栖菜多酚对RAW264.7细胞的安全质量浓度范围为0～160 μg/mL。与LPS组相比,羊栖菜多酚剂量依赖性降低巨噬细胞吞噬能力并抑制NO的生成。同时,羊栖菜多酚下调促炎细胞因子（IL-1β、IL-6、TNF-α）和炎症诱导酶（iNOS、COX-2）的mRNA水平,且作用效果与给药剂量及LPS刺激时间相关。这些炎性介质的表达与羊栖菜多酚抑制p38 MAPK和NF-κB p65的激活有关。结论：羊栖菜多酚可通过减弱p38 MAPK和NF-κB p65信号通路的激活水平,抑制下游炎症介质的转录表达,从而缓解LPS诱导的巨噬细胞炎症反应。     

DPA体外抗炎活性及机制研究

以脂多糖(LPS)诱导的RAW264.7巨噬细胞为炎症的体外模型,以炎症介质【NO和前列腺素E2(PGE2)】和炎症因子(TNF-α,IL-1β,IL-6和IL-10)为指标,研究二十二碳五烯酸(DPA)的体外抗炎活性,并从NF-κB代谢通路的角度探究DPA体外抗炎活性的作用机制。结果表明,DPA可以显著抑制iNOS和COX-2的表达,抑制炎症介质NO和PGE2的分泌,并且,DPA通过调控促炎因子(TNF-α,IL-1β,IL-6)和抑炎因子(IL-10)的平衡发挥抗炎作用。Western blot结果表明,DPA显著抑制NF-κB代谢通路中p50和p65的磷酸化,限制p50/p65核内转移,抑制炎症级联反应,发挥抗炎作用。     

基于LPS诱导RAW264.7巨噬细胞炎症模型的榴莲壳多酚抗炎作用及其分子机制

采用80%甲醇溶剂提取榴莲壳中多酚组分,并测定其总酚和总黄酮含量,基于体外化学方法和LPS诱导的RAW 264.7巨噬细胞炎症模型,探究榴莲壳的抗氧化和抗炎活性及其分子机制。结果表明,榴莲壳提取物富含多酚类化合物,具有较强的体外抗氧化活性,且在LPS诱导的RAW 264.7巨噬细胞炎症模型中能降低一氧化氮合酶(iNOS)和环氧合酶-2(COX-2)的表达,减少NO和ROS的产生,并降低炎症细胞因子(TNF-α、IL-1β和IL-6)在基因和蛋白水平上的表达,从而展现较强的抗炎活性。其内在的分子机制可能是通过抑制IκB-α和p65蛋白磷酸化,降低NF-κB信号通路的表达,减少机体的炎症损伤。     

沙葱总黄酮水洗组分的体外抗炎活性

本实验在体外应用脂多糖（lipopolysaccharides,LPS）诱导小鼠腹腔巨噬细胞建立炎症模型的基础上,探讨沙葱总黄酮水洗组分的体外抗炎活性。应用CCK-8法检测0、50、100、200、400、800 μg/mL沙葱总黄酮水洗组分对小鼠腹腔巨噬细胞增殖活力的影响;将细胞分为空白对照组、LPS应激模型组及不同质量浓度沙葱总黄酮水洗组分预处理组,采用Griess法及酶联免疫吸附测定法分别测定各处理组细胞上清液中NO浓度和肿瘤坏死因子-α（tumor necrosis factor-α,TNF-α）、白细胞介素（interleukin,IL）-1β、IL-6、IL-10的质量浓度,反转录实时荧光定量聚合酶链式反应法检测各处理组细胞中诱导型一氧化氮合酶（inducible nitric oxide synthase,iNOS）、TNF-α、IL-1β、IL-6、IL-10、髓样分化蛋白（myeloid differential protein,MyD）88、核因子 κB（nuclear factor κB,NF-κB）mRNA的表达水平。结果显示：与对照组相比,沙葱总黄酮水洗组分在50～800 μg/mL时对小鼠腹腔巨噬细胞无明显细胞毒性作用。与LPS应激模型组相比,沙葱总黄酮水洗组分能够极显著抑制促炎介质NO、TNF-α、IL-1β、IL-6质量浓度及其mRNA的表达（P＜0.01或P＜0.001）;高度显著提高抗炎细胞因子IL-10质量浓度及其mRNA的表达（P＜0.001）,且呈剂量依赖效应;极显著降低MyD88、NF-κB mRNA的表达水平（P＜0.01或P＜0.001）。由此得出,沙葱总黄酮水洗组分对LPS诱导的小鼠腹腔巨噬细胞具有抗炎作用,其抗炎活性可能是通过抑制促炎性介质NO、TNF-α、IL-1β、IL-6的分泌并提高抗炎性细胞因子IL-10的质量浓度实现的,其作用机制可能与NF-κB信号通路有关。     

高核苷酸酵母水解物对脂多糖诱导RAW264.7细胞免疫调节的影响

为探讨高核苷酸酵母水解物对RAW264.7小鼠巨噬细胞免疫活性调节及其相关作用机制,采用脂多糖(1μg/mL)刺激RAW264.7细胞来建立体外细胞炎症模型,以不同质量浓度的高核苷酸酵母水解物干预来明确其抗炎效果。ELISA法检测细胞培养上清中白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)、白细胞介素-1β(IL-1β)的含量,RT-PCR法检测相关mRNA表达水平,Western blot法检测细胞iNOS及胞核内核转录因子NF-κBp65蛋白水平。结果表明:高核苷酸酵母水解物作用RAW264.7细胞的安全范围≤150μg/mL。与LPS模型组相比,质量浓度60～150μg/mL的高核苷酸酵母水解物能显著增强RAW264.7吞噬能力,高核苷酸酵母水解物(60～150μg/mL)能有效抑制脂多糖(lipopolysaccharide,LPS)诱导RAW264.7细胞释放NO、TNF-α、IL-1β和IL-6炎症因子及相关mRNA表达,降低iNOS及NF-κB蛋白水平。研究结果证实了高核苷酸酵母水解物对LPS刺激RAW264.7细胞炎症的保护作用,作用机制与NF-κB通路有关,为食疗干预慢性病提供一定的理论依据。     

竹节参总提物对LPS诱导BV2小胶质细胞炎症反应的保护作用

本文研究了竹节参总提物对脂多糖（lipopolysaccharide, LPS）刺激所致的BV2小胶质细胞炎症反应的保护作用及其机制。MTT筛选不同浓度竹节参总提取对小胶质细胞增殖影响;一氧化氮试剂盒检测NO表达水平;PCR检测IL-1β、TNF-α和iNOS的mRNA表达;免疫荧光检测NF-κB表达及定位。竹节参总提物在3.125~50 mg/L对正常状态及LPS 1 μg/mL诱导的炎症状态下小胶质细胞生长均无明显影响;与正常组比较,模型组NO释放量是其11.82倍,TNF-α、IL-1β和iNOS的mRNA表达量分别是正常组的1.35、1.78和2.50倍,并且NF-κB核移位明显;与模型组比较,竹节参总提物在3.125~50 mg/L均能不同程度抑制NO释放量,并减少IL-1β、TNF-α和iNOS的mRNA表达水平;25、50 mg/L竹节参总提物有效抑制了小胶质细胞中NF-κB的核移位。竹节参总提物可减轻LPS诱导的BV2小胶质细胞炎症反应,其主要机制为抑制NF-κB的核移位,进而降低了炎症因子的分泌。     

野生蒙古口蘑多糖通过STAT3和HIF-1α通路对LPS诱导的巨噬细胞发挥抗炎调节作用

研究蒙古口蘑多糖提取物对脂多糖(LPS)诱导小鼠单核巨噬细胞(RAW264.7细胞)炎症保护作用的机理。用不同预量质量浓度的蒙古口蘑多糖处理RAW264.7细胞,通过CCK-8法检测细胞活性;Griess法测定NO的产生量;酶联免疫吸附测定肿瘤坏死因子-α(TNF-α),白介素1β(IL-1β);实时定量检测缺氧诱导因子1α(HIF-1α)、诱导型一氧化氮和酶(iNOS)、核因子κB(NF-κB)、信号转导和转录激活因子3(STAT3)和环氧化酶2(COX-2)的mRNA表达量;免疫印迹检测HIF-1α、NF-κB、STAT3和COX-2蛋白质的表达量。结果表明,蒙古口蘑多糖提取物作用RAW264.7细胞的最大剂量为100μg/mL;与LPS模型组相比,蒙古口蘑多糖提取物质量浓度0.01、0.1、1、10μg/mL均能有效抑制NO(抑制率分别是38.21%、64.17%、58.16%和79.42%)、TNF-α(抑制率分别为27.91%、36.31%、36.14%、45.09%)和IL-1β(抑制率分别为47.75%、58.47%、60.21%、64.55%)的产生;在质量浓度为10μg/mL的蒙古口蘑多糖提取物作用下,除NF-κB外,HIF-1α、STAT3、COX-2和iNOS的mRNA的表达量均降低;免疫印迹的结果也显示,除NF-κB外,HIF-1α、NF-κB、STAT3及COX-2的蛋白质的表达量均有不同程度的降低。蒙古口蘑多糖提取物对脂多糖(LPS)诱导的RAW264.7细胞具有保护作用,其抗炎的机制主要通过JAK-STAT3及HIF-1α信号通路,而不主要通过NF-κB信号通路发挥作用。     

黑灵芝多糖体内抗炎活性及对甘露糖受体表达的影响

目的:研究水溶性黑灵芝多糖(a water-soluble polysaccharides from Ganoderma atrum,PSG)-1体内抗炎活性及对甘露糖受体(mannose receptor,MR)表达的影响。方法:腹腔注射脂多糖(lipopolysaccharide,LPS)建立小鼠体内炎症模型,随机分为5组:正常对照组、LPS组、PSG-1(高、中、低剂量,分别为100、50、25 mg/(kg·d))+LPS组。流式细胞仪检测巨噬细胞中MR表达、吞噬功能及活性氧(reactive oxygen species,ROS)生成,用酶联免疫吸附测定(enzyme-linked immunosorbent assay,ELISA)分析血清中肿瘤坏死因子(tumor necrosis factor,TNF)-α、白细胞介素(interleukin,IL)-1β和IL-6的含量。结果:与正常对照组相比,LPS组腹腔巨噬细胞表面MR表达量下降,与LPS组相比,PSG-1+LPS组腹腔巨噬细胞表面MR表达量极显著增加(P0.01);与正常对照组相比,LPS组腹腔巨噬细胞吞噬能力和ROS生成增加,与LPS组相比,PSG-1+LPS组腹腔巨噬细胞吞噬功能和ROS生成显著降低(P0.05,P0.0 1);与正常对照组相比,小鼠经LPS处理后,血清中TNF-α、IL-1β和I L-6的分泌水平极显著升高(P0.01);与LPS组相比,小鼠灌胃PSG-1后,PSG-1(高剂量)+LPS组中TNF-α的含量显著降低(P0.05),IL-1β和IL-6的分泌水平无显著差异。结论:PSG-1具有抗炎作用,可部分抑制LPS诱导的体内炎症,其机理与PSG-1促进小鼠腹腔巨噬细胞表面MR的表达、抑制巨噬细胞向M1型极化有关。     

Comparative effects of tocotrienol-rich fraction, α-tocopherol and α-tocopheryl acetate on inflammatory mediators and nuclear factor kappa B expression in mouse peritoneal macrophages

The effects of tocotrienol-rich fraction (TRF), α-tocopherol (T) and α-tocopheryl acetate (TA) on lipopolysaccharide (LPS)-induced inflammatory responses in mouse peritoneal macrophages were examined. Results showed that at 5–30 μg/ml, all test compounds plus 1 μg/ml LPS exhibited no cytotoxic effects on macrophage cells. Compared with T and TA, TRF showed the strongest anti-inflammatory activity as demonstrated by its potency in inhibiting the LPS-induced nitric oxide (NO), prostaglandin E₂ (PGE₂), and proinflammatory cytokine (TNF-α, IFN-γ, IL-1β and IL-6) production. At 10 μg/ml, it significantly blocked the LPS induction of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) expression, but has no effect on cyclooxygenase-1 (COX-1). Furthermore, TRF also showed a greater inhibition on the nuclear factor kappa B (NF-κB) expression than T and TA. These results suggest that TRF could be a better agent than T and TA for use in the prevention of chronic inflammatory diseases.     

Anti-inflammatory activity of ethanolic extract of Sargassum sagamianum in RAW 264.7 cells

Aim of the study is to evaluate the antiinflammatory effects of ethanolic extract of the marine brown alga Sargassum sagamianum collected from Yeonhwari coast of Korea. Ethanolic extract of S. sagamianum (SA-E extract) inhibited expression of nitric oxide (NO) and cytokines (IL-6, IL-1β, and TNF-α) as well as inducible nitric oxide synthase (iNOS) and cyclooxygenase (COX)-2 in lipopolysaccharide (LPS)-induced RAW 264.7 cells without affecting cell viability. In addition, the expression of nuclear factor (NF)-κB p65 was suppressed by SA-E extract. Furthermore, the rate of formation of edema in the mouse ear was reduced by 46% at the highest dose tested (250 mg/kg) compared to that in the control. This study suggests that SA-E extract exerts potent inhibitory effects on LPS-induced expression of inflammatory mediators such as NO, iNOS, COX-2, and cytokines in macrophages through suppression of the NF-κB p65 pathway. SA-E extract might have potential clinical applications as an anti-inflammatory agent.     

Anti-inflammatory effect of flavonoids isolated from Korea Citrus aurantium L. on lipopolysaccharide-induced mouse macrophage RAW 264.7 cells by blocking of nuclear factor-kappa B (NF-κB) and mitogen-activated protein kinase (MAPK) signalling pathways

Citrus fruits are an abundant source of various flavonoids, which have been used as a traditional herbal medicine in Korea and China. Most flavonoids are known to have anti-oxidant, anti-bacterial and analgesic properties. In this study, it was examined whether flavonoids (nobiletin, naringin and hesperidin) isolated from Korea Citrus aurantium L. inhibited the pro-inflammatory mediators including cytokines by blocking nuclear factor-kappa B (NF-κB) and mitogen-activated protein kinase (MAPK) signalling in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. The flavonoids suppressed mRNA and protein expression of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) in LPS-induced macrophages. The molecular mechanism was associated with inhibition of the degradation/phosphorylation of I-κB-α and nuclear translocation of the NF-κB p-65 as well as phosphorylation of MAPK by flavonoids. These results suggest that flavonoids have anti-inflammatory effects by suppressing expression of COX-2, iNOS and cytokines by blocking the NF-κB and MAPK signalling in RAW 264.7 macrophages.     

苦荞蛋白源肽AFYRW对脂多糖诱导的人脐静脉内皮细胞损伤的影响

目的：探讨苦荞蛋白源肽AFYRW在脂多糖（lipopolysaccharide,LPS）诱导的血管内皮细胞损伤中的作用。方法：采用LPS诱导人脐静脉内皮细胞（human umbilical vein endothelial cells,HUVECs）建立血管内皮细胞炎症损伤模型;采用CCK8试剂盒检测AFYRW对细胞增殖活力的影响;Western blot检测血管细胞黏附分子-1(vascular cell adhension molecule-1,VCAM-1)、细胞内黏附分子-1(intercellular adhension molecule-1,ICAM-1)、白细胞介素（interleukin,IL）-6、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、内皮型一氧化氮合酶（endothelial nitric oxide synthase,e NOS）、诱导型一氧化氮合酶（inducible nitricoxide synthase,iNOS）水平及核因子-κB(nuclear factor kappa-B,NF-κB)p65的磷酸化水平;酶法...